内生型转铁蛋白-金纳米簇探针的合成及其对前列腺癌的靶向成像研究

目的 以转铁蛋白(transferrin,Tf)为模板,氯金酸为原料,原位合成靶向荧光探针-内生型转铁蛋白-金纳米簇(gold nanoclusters,AuNCs),并以前列腺癌(prostate cancer,PCa)为模型,探讨该荧光探针的靶向成像效果.方法 利用场发射透射电子显微镜及粒度分析仪分析所合成的荧光探针的形貌、水合粒径及分布;利用紫外-可见分光光度计及荧光分光光度计对其光学性能进行表征;MTT法检测细胞毒性;并通过细胞荧光成像及竞争抑制实验表征其肿瘤的特异性靶向效果;静脉注射Tf-AuNCs至PC-3种植瘤鼠体内,进行连续荧光成像,评价其肿瘤靶向成像效果;通过组织病理学明确其活体毒性.结果 本研究制备的Tf-AuNCs粒径约为3 nm,荧光发射峰为700 nm,保留了Tf及金纳米簇的性能;细胞成像结果显示Tf-AuNCs对前列腺肿瘤细胞(PC-3)特异性靶向效果好;小动物活体成像结果显示Tf-AuNCs探针仅需15 min即可准确显示肿瘤部位,至2h时肿瘤区荧光强度达峰值,表明其具有良好的活体肿瘤成像能力;病理学结果表明Tf-AuNCs具有良好的生物相容性.结论 Tf-AuNCs荧光探针具有良好的光学性能、特异靶向能力,并具有优异的荧光成像能力及良好的生物相容性,将有望应用于PCa的早期影像学分析.     

Assessment of oxidative stress induced by gold nanorods following intra-tracheal instillation in rats

Gold nanorods (GNRs) are used for their wide variety of applications in various industries. There is a little availability of data related to toxicity and ecological implications of these GNRs. The study evaluated the oxidative stress induction following intra-tracheal instillation of 1 and 5?mg/kg b.w. doses of 10 and 25?nm GNRs by estimating various oxidative stress markers including lipid peroxidation (malondialdehyde; MDA), glutathione (GSH), superoxide dismutase (SOD), catalase and total antioxidant capacity (TAC) after 1?day, 1?week, 1?month, and 3?months post exposure periods. The results have shown increased MDA levels and decreased GSH levels following 1?day and 1?week post exposure periods, indicating induction of oxidative stress. Also, the SOD, catalase and TAC levels were significantly decreased following exposure of both 10 and 25?nm GNRs after 1?day and 1?week after exposures, indicating the inhibition of antioxidant defense mechanisms. Moreover, the 10?nm GNRs at 5?mg/kg dose displayed greater changes in all the estimated parameters, representing dose and size based induction of oxidative stress by GNRs. In contrast, a little change was observed during 1?month and 3?months post exposure periods, may be due to recovery. Finally, the GNRs induced dose-size-dependent oxidative stress induction by various oxidative stress markers following intra-tracheal instillation in rats.     

Gold nanoparticles: Distribution,bioaccumulation and toxicity. In vitro and in vivo studies

Concerns about the bioaccumulation and toxicity of gold nanoparticles inside humans have recently risen. HT-29 and HepG2 cell lines and Wistar rats were exposed to 10, 30 or 60 nm gold nanoparticles to determine their tissue distribution, subcellular location and deleterious effects. Cell viability, ROS production and DNA damage were evaluated in vitro. Lipid peroxidation and protein carbonylation were determined in liver. ICP-MS measurements showed the presence of gold in intestine, kidney, liver, spleen, feces and urine. Subcellular locations of gold nanoparticles were observed in colon cells and liver samples by transmission electron microscopy. Inflammatory markers in liver and biochemical parameters in plasma were measured to assess the inflammatory status and presence of tissue damage. The size of the nanoparticles determined differences in the biodistribution and the excretion route. The smallest nanoparticles showed more deleterious effects, confirmed by their location inside the cell nucleus and the higher DNA damage.     

Endoscopic molecular imaging of early gastric cancer using fluorescently labeled human H-ferritin nanoparticle

Optical imaging technologies improve clinical diagnostic accuracy of early gastric cancer (EGC). However, there was a lack of imaging agents exhibiting molecular specificity for EGCs. Here, we employed the dye labeled human heavy-chain ferritin (HFn) as imaging nanoprobe, which recognizes tumor biomarker transferrin receptor 1 (TfR1), to enable specific EGC imaging using confocal laser endomicroscopy (CLE). TfR1 expression was initially examined in vitro in gastric tumor cells and in vivo through whole-body fluorescence and CLE imaging in tumor-bearing mice. Subsequently, dye labeled HFn was topically applied to resected human tissues for EGC detection. CLE analysis of TfR1-targeted fluorescence imaging allowed distinction of neoplastic from non-neoplastic tissues (P?<?0.0001), and TfR1 expression level was found to correlate with EGC differentiation degrees (P?<?0.0001). Notably, the CLE evaluation correlated well with the immunohistochemical findings (κ-coefficient: 0.8023). Our HFn-nanoprobe-based CLE increases the accuracy of EGC detection and enables visualization of tumor margins and endoscopic resection.     

Nano in nano: Biosynthesized gold and iron nanoclusters cargo neoplastic exosomes for cancer status biomarking

Timely detection is crucial for successful treatment of cancer. The current study describes a new approach that involves utilization of the tumor cell environment for bioimaging with in-situ biosynthesized nanoscale gold and iron probes and subsequent dissemination of Au-Fe nanoclusters from cargo exosomes within the circulatory system. We have isolated the Au-Fe cargo exosomes from the blood of the treated murine models after in situ biosyntheses from their respective pre-ionic solutions (HAuCl₄, FeCl₂), whereas Na₂SeO₃ supplementation added into Au lethal effect. The microarray data of various differentially expressed genes revealed the up-regulated tumor ablation and metal binding genes in SGC-7901 cell lines after treatment with Au-Fe-Se triplet ionic solution. The isolation of Au-Fe nanoclusters cargo exosomes (nano in nano) after secretion from deeply seated tumors may help in early diagnosis and reveal the tumor ablation status during and after the relevant treatment like radio-chemo therapies et al.     

Neuroprotective effect of gold nanoparticles composites in Parkinson's disease model

Parkinson’s disease (PD) is second most common neurodegenerative disorder worldwide. Although drugs and surgery can relieve the symptoms of PD, these therapies are incapable of fundamentally treating the disease. For PD patients, over-expression of α-synuclein (SNCA) leads to the death of dopaminergic neurons. This process can be prevented by suppressing SNCA over-expression through RNA interference. Here, we successfully synthesized gold nanoparticles (GNP) composites (CTS@GNP-pDNA-NGF) via the combination of electrostatic adsorption and photochemical immobilization, which could load plasmid DNA (pDNA) and target specific cell types. GNP was transfected into cells via endocytosis to inhibiting the apoptosis of PC12 cells and dopaminergic neurons. Simultaneously, GNP composites are also used in PD models in vivo, and it can successfully cross the blood-brain barrier by contents of GNP in the mice brain. In general, all the works demonstrated that GNP composites have good therapeutic effects for PD models in vitro and in vivo.     

Targeted-gene silencing of BRAF to interrupt BRAF/MEK/ERK pathway synergized photothermal therapeutics for melanoma using a novel FA-GNR-siBRAF nanosystem

Melanoma is significantly associated with mutant BRAF gene, a suitable target for siRNA-based anti-melanoma therapy. However, a tumor-specific delivery system is a major hurdle for clinical applications. Here, we developed a novel nano-carrier, FA-GNR-siBRAF for safe topical application, which consists of folic acid (FA) as the tumor-targeting moiety, golden nanorods (GNR) providing photothermal capability to kill tumor cells under laser irradiation, and siRNA specifically silencing BRAF (siBRAF). The in vitro and in vivo results revealed that FA-GNR-siBRAF displayed high transfection rates, and subsequently induced remarkable gene knockdown of BRAF, resulting in suppression of melanoma growth due to the interruption of the MEK/ERK pathway. Combinatorial photothermal effects and BRAF knockdown by FA-GNR-siBRAF effectively killed tumor cells through apoptosis, with enhanced efficiency than individual treatments. Therefore, the FA-GNR-siBRAF simultaneously induced BRAF gene silencing and photothermal effects which achieved synergistic efficacy in the treatment of melanoma, paving a new path for developing clinical treatment methods for melanoma.     

Development of a novel quantitative real‐time PCR assay with lyophilized powder reagent to detect African swine fever virus in blood samples of domestic pigs in China

African swine fever (ASF) is a devastating disease, which is causing huge economic losses in China. Therefore, it is urgent to provide a rapid, highly specific and sensitive diagnostic method for the detection of African swine fever virus (ASFV), the ASF infectious agent. In this study, a novel quantitative real‐time polymerase chain reaction (qPCR) assay with lyophilized powder reagents (LPR), targeting the major structural protein p72 gene, was established for the detection of ASFV. This assay had many advantages, such as saving time and money, good sensitivity and repeatability. The sensitivity of this assay was 100 copies/μl of ASFV plasmid templates, and the assay showed 10‐fold greater sensitivity than a qPCR assay recommended by OIE. Furthermore, specificity analysis showed that qPCR with LPR for ASFV had no cross‐reactivity with other important swine pathogens. In clinical diagnoses of 218 blood samples of domestic pigs in China, the positive rate of the diagnosis of ASFV by qPCR with the LPR and commercial kit reached 80.73% (176/218) and 76.61% (167/218) respectively. The coincidence rate between the two assays is 92.20% (201/218), and kappa value is 0.768 (p < .0001) by SPSS analysis. The overall agreement between the two assays was 95.87% (209/218). Further Pearson correlation and linear regression analysis showed a significant correlation between the two assays with an R² value of 0.9438. The entire procedure, from specimen processing to result reporting, can be completed within 2 hr. Our results demonstrated that the qPCR‐LPR assay is a good laboratory diagnostic tool for sensitive and efficient detection of ASFV.