Lyophilization Based Isolation of Exosomes.

Exosomes are nanoscale extracellular vesicles which regulate intercellular communication. They have great potential for application in nanomedicine. However, techniques for their isolation are limited by requirements for advanced instruments and costly reagents. In this study, we developed a lyophilization-based method for isolating exosomes from cultured cells. The isolated exosomes were characterized for protein content using Bradford assay, and for size distribution and shape using scanning electron microscopy (SEM) and nanoparticles tracking analysis (NTA). In addition, CD63, CD9, CD81, HSP70 and TSG101 were evaluated as essential exosomal surface markers using Western blot. Drug loading and release studies were performed to confirm their drug delivery properties using an in vitro model. Exosomes were also loaded with commercial dyes (Cy5, Eosin) for the evaluation of their drug delivery properties. All these characterizations confirmed successful exosome isolation with measurements of less than 150 nm, having a typical shape, and by expressing the known exosome surface protein markers. Finally, tyrosine kinase inhibitors (dasatinib and ponatinib) were loaded on the exosomes to evaluate their anticancer effects on leukemia cells (K562 and engineered Ba/F3-BCR-ABL) using MTT and Annexin-PI assays. The expression of MUC1 protein on the exosomes isolated from MCF-7 cells also indicated that their potential diagnostic properties were intact. In conclusion, we developed a new method for exosome isolation from cultured cells. These exosomes met all the essential requirements in terms of characterization, drug loading and release ability, and inhibition of proliferation and apoptosis induction in Ph+ leukemia cells. Based on these results, we are confident in presenting the lyophilization-based exosome isolation method as an alternative to traditional techniques for exosome isolation from cultured cells.

In Vivo Biosynthesized Zinc and Iron Oxide Nanoclusters for High Spatiotemporal Dual-Modality Bioimaging of Alzheimer's Disease.

Alzheimer's disease is still incurable and neurodegenerative, and there is a lack of detection methods with high sensitivity and specificity. In this study, by taking different month old Alzheimer's mice as models, we have explored the possibility of the target bioimaging of diseased sites through the initial injection of zinc gluconate solution into Alzheimer's model mice post-tail vein and then the combination of another injection of ferrous chloride (FeCl2) solution into the same Alzheimer's model mice post-stomach. Our observations indicate that both zinc gluconate solution and FeCl2 solution could cross the blood-brain barrier (BBB) to biosynthesize the fluorescent zinc oxide nanoclusters and magnetic iron oxide nanoclusters, respectively, in the lesion areas of the AD model mice, thus enabling high spatiotemporal dual-modality bioimaging (i.e., including fluorescence bioimaging (FL) and magnetic resonance imaging (MRI)) of Alzheimer's disease for the first time. The result presents a novel promising strategy for the rapid and early diagnosis of Alzheimer's disease.

Biosynthesized Gold Nanoclusters and Iron Complexes as Scaffolds for Multimodal Cancer Bioimaging.

Cancer treatment has a far greater chance of success if the neoplasm is diagnosed before the onset of metastasis to vital organs. Hence, cancer early diagnosis is extremely important and remains a major challenge in modern therapeutics. In this contribution, facile and new method for rapid multimodal tumor bioimaging is reported by using biosynthesized iron complexes and gold nanoclusters via simple introduction of AuCl4- and Fe2+ ions. The observations demonstrate that the biosynthesized Au nanoclusters may act as fluorescent and computed tomography probes for cancer bioimaging while the iron complexes behave as effective contrast agent for magnetic resonance imaging. The biosynthesized iron complexes and gold nanoclusters are found biocompatible in vitro (MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay) and in vivo for all the vital organs of circulatory and excretory system. These observations raise the possibility that the biosynthesized probes may find applications in future clinical diagnosis for deep seated early neoplasms by multimodal imaging.

Iguratimod: a valuable remedy from the Asia Pacific region for ameliorating autoimmune diseases and protecting bone physiology.

Autoimmune diseases are affected by complex pathophysiology involving several cell types, cytokines, antibodies, and mimicking factors. Different drugs are used to ameliorate these autoimmune reactions, including nonsteroidal anti-inflammatory drugs (NSAIDs), corticosteroids, antiantibodies, and small molecular drugs (DMARDs), and they are clinically in vogue for diseases such as rheumatoid arthritis (RA). Nevertheless, low cost-effectiveness, reduced efficacy, adverse effects, and patient nonresponse are unappealing factors driving the development of new drugs such as iguratimod. Iguratimod is primarily used to ameliorate RA in Japanese and Chinese clinics. However, its efficacy against other autoimmune ailments is also under intense investigation, and the number of investigations is becoming increasingly larger with each passing day. The articular structure comprises synovium, ligaments, and bone. The latter is more complex than the others since it regulates blood cells and autoimmunity in addition to providing skeletal support to the body. Therefore, its protection is also of prime importance in RA and other autoimmune diseases. Herein, we have highlighted the role of iguratimod in autoimmune diseases and bone protection. We suggest that iguratimod's unique mode of action compared with that of other DMARDs and its good patient response makes it a suitable antirheumatic and bone-protecting drug.

Novel multifunctional nanospheres of Zn1/3Fe8/3O4@Ag: synthesis, properties and application for multi-modality tumor imaging.

New multifunctional nanospheres have been designed and synthesized through a green and facile strategy, which could be readily used in multi-modality tumor imaging through near-infrared fluorescence (NIRF) imaging, magnetic resonance imaging (MRI) and computed tomography (CT) imaging. Such nanospheres are made of porous superparamagnetic Zn1/3Fe8/3O4 nanosphere cores covered by a thin layer shell of Ag clusters. While the shell of Ag clusters provides efficient NIRF imaging, the Zn1/3Fe8/3O4 nanosphere cores allow external magnetic manipulation and readily facilitates their utilization for MRI and CT imaging.

Protective effect of TiO2 nanowhiskers on Tetra Sulphonatophenyl Porphyrin (TSPP) complexes induced oxidative stress during photodynamic therapy.

BACKGROUND: Tetra Sulphonatophenyl Porphyrin (TSPP) is well known photosensitizer for photodynamic therapy; nevertheless, its well-known adverse effects hamper its potential use. Recently, nano TiO2's potential role in biomedical has been defined for various disease theranostics, including cancer and other infections. Thus, in this contribution we have explored the possibility of utilizing TiO2 nanowhiskers as novel strategy to lower TSPP adverse effects both in vitro, and in vivo. METHODS: Various concentrations of TSPP, TiO2-TSPP, and TiO2 were injected to three different rat groups, while fourth group was kept as control. Toxic effects were evaluated on excretory and circulatory system by using histopathology, fluorescent microscopy, complete blood cells count (CBC) and serum enzymes. RESULTS: In complete blood cells count, all cells were significantly (p<0.01) affected by the various concentration and treatment groups. The various dose concentrations and treatment also significantly (p<0.01) affected the serum enzyme parameters including AST, ALT, LDH, Creatinine and BUN level. The low concentration of TSPP-TiO2 was found to be the safest, on the bases of serum enzyme parameters, CBC, histopathology, and fluorescent microscopic analysis. The MTT assay was used to evaluate in vitro cytotoxicity, and the results demonstrated maximum viability in illuminated TSPP-TiO2 nanowhiskers group when compared with TSPP treated group. CONCLUSIONS: It was evident that increase in concentration of TSPP increased the toxic effects; however, the TiO2 nanowhiskers combination with TSPP decreased these adverse effects. Moreover, TSPP (0.1 mM) combined with TiO2 nanowhiskers (0.6 mM) was safer than TSPP (0.1 mM) alone.

Recent advances in nano scaffolds for bone repair.

Biomedical applications of nanomaterials are exponentially increasing every year due to analogy to various cell receptors, ligands, structural proteins, and genetic materials (that is, DNA). In bone tissue, nanoscale materials can provide scaffold for excellent tissue repair via mechanical stimulation, releasing of various loaded drugs and mediators, 3D scaffold for cell growth and differentiation of bone marrow stem cells to osteocytes. This review will therefore highlight recent advancements on tissue and nanoscale materials interaction.

In vivo target bio-imaging of Alzheimer's disease by fluorescent zinc oxide nanoclusters.

Alzheimer's disease (AD) is an irreversible neurodegenerative disease which is difficult to cure. When Alzheimer's disease occurs, the level of zinc ions in the brain changes, and the relevant amount of zinc ions continue decreasing in the cerebrospinal fluid and plasma of Alzheimer's patients with disease exacerbation. In view of these considerations, we have explored a new strategy for the in vivo rapid fluorescence imaging of Alzheimer's disease through target bio-labeling of zinc oxide nanoclusters which were biosynthesized in vivo in the Alzheimer's brain via intravenous injection of zinc gluconate solution. By using three-month-old and six-month-old Alzheimer's model mice as models, our observations demonstrate that biocompatible zinc ions could pass through the blood-brain barrier of the Alzheimer's disease mice and generate fluorescent zinc oxide nanoclusters (ZnO NCs) through biosynthesis, and then the bio-synthesized ZnO NCs could readily accumulate in situ on the hippocampus specific region for the in vivo fluorescent labeling of the affected sites. This study provides a new way for the rapid diagnosis of Alzheimer's disease and may have promising prospects in the effective diagnosis of Alzheimer's disease.

In Situ Biosynthesis of Fluorescent Platinum Nanoclusters: Toward Self-Bioimaging-Guided Cancer Theranostics.

Among the noble-metal clusters, very few reports about platinum clusters were used as bioimaging probes of tumors except as a reducing catalyst. It is first established herein that the biocompatible platinum nanoclusters are spontaneously biosynthesized by cancerous cells (i.e., HepG2 (human hepatocarcinoma), A549 (lung cancer), and others) rather than noncancerous cells (i.e., L02 (human embryo liver cells)) when incubated with micromolar chloroplatinic acid solutions. These in situ biosynthesized platinum nanoclusters could be readily realized in a biological environment and emit a bright fluorescence at 460 nm, which could be further utilized to facilitate an excellent cancer-cell-killing efficiency when combined with porphyrin derivatives for photothermal treatment. This raises the possibility of providing a promising and precise bioimaging strategy for specific fluorescent self-biomarking of tumor locations and realizing fluorescence imaging-guided photothermal therapy of tumors.

Bio-imaging and Photodynamic Therapy with Tetra Sulphonatophenyl Porphyrin (TSPP)-TiO2 Nanowhiskers: New Approaches in Rheumatoid Arthritis Theranostics.

Since Rheumatoid arthritis (RA) is one of the major human joint diseases with unknown etiology, the early diagnosis and treatment of RA remains a challenge. In this contribution we have explored the possibility to utilize novel nanocomposites of tetera suplhonatophenyl porphyrin (TSPP) with titanium dioxide (TiO2) nanowhiskers (TP) as effective bio-imaging and photodynamic therapeutic (PDT) agent for RA theranostics. Our observations demonstrate that TP solution PDT have an ameliorating effect on the RA by decreasing significantly the IL-17 and TNF-α level in blood serum and fluorescent imaging could enable us to diagnose the disease in subclinical stages and bio-mark the RA insulted joint.