Deficiency of the dual ubiquitin/SUMO ligase Topors results in genetic instability and an increased rate of malignancy in mice

<p>Abstract</p> <p>Background</p> <p>Topors is a nuclear protein that co-localizes with promyelocytic leukemia bodies and has both ubiquitin and SUMO E3 ligase activity. Expression studies implicated Topors as a tumor suppressor in various malignancies. To gain insight into the function of Topors, we generated a Topors-deficient mouse strain.</p> <p>Results</p> <p>Mice homozygous for a mutant Topors allele exhibited a high rate of perinatal mortality and decreased lifespan. In addition, heterozygotes were found to have an increased incidence of malignancy, involving a variety of tissues. Consistent with this finding, primary embryonic fibroblasts lacking Topors exhibited an increased rate of malignant transformation, associated with aneuploidy and defective chromosomal segregation. While loss of Topors did not alter sensitivity to DNA-damaging or microtubule-targeting agents, cells lacking Topors exhibited altered pericentric heterochromatin, manifested by mislocalization of HP1α and an increase in transcription from pericentric major satellite DNA. Topors-deficient cells exhibited a transcriptional profile similar to that of cells treated with histone deacetylase inhibitors, and were resistant to the anti-proliferative effects of the histone deacetylase inhibitor trichostatin A.</p> <p>Conclusion</p> <p>These results indicate a unique role for Topors in the maintenance of genomic stability and pericentric heterochromatin, as well as in cellular sensitivity to histone deacetylase inhibitors.</p

Quantum dot/antibody conjugates for in vivo cytometric imaging in mice

Multiplexed, phenotypic, intravital cytometric imaging requires novel fluorophore conjugates that have an appropriate size for long circulation and diffusion and show virtually no nonspecific binding to cells/serum while binding to cells of interest with high specificity. In addition, these conjugates must be stable and maintain a high quantum yield in the in vivo environments. Here, we show that this can be achieved using compact (~15 nm in hydrodynamic diameter) and biocompatible quantum dot (QD) -Ab conjugates. We developed these conjugates by coupling whole mAbs to QDs coated with norbornene-displaying polyimidazole ligands using tetrazine–norbornene cycloaddition. Our QD immunoconstructs were used for in vivo single-cell labeling in bone marrow. The intravital imaging studies using a chronic calvarial bone window showed that our QD-Ab conjugates diffuse into the entire bone marrow and efficiently label single cells belonging to rare populations of hematopoietic stem and progenitor cells (Sca1[superscript +]c-Kit[superscript +] cells). This in vivo cytometric technique may be useful in a wide range of structural and functional imaging to study the interactions between cells and between a cell and its environment in intact and diseased tissues.National Institutes of Health (U.S.) (Grant U54-CA151884)National Institutes of Health (U.S.) (Grant P41-EB015871-26A1)Samsung Scholarship Foundation (Graduate Student Fellowship)Massachusetts Institute of Technology. Institute for Soldier Nanotechnologies (Grant W911NF-07-D-0004

Development of Gold-Based Phototheranostic Nanoagents through a Bioinspired Route and Their Applications in Photodynamic Therapy

Green synthesis of nanoparticles using phytoconstituents has been widely accepted. However, further applications for delivery, diagnosis, and therapy are not yet fully established. Thus, bioinspired nanomedicinal diagnostic agents have been developed with antioxidant, diagnostic, as well as therapeutic properties. The sustainable biological synthesis of fluorescent gold nanoparticles using <i>Syzygium cumini</i> fruit extract presents a simplified process for the production of surface-functionalized nanoparticles with good antioxidant potential. The gold-based phototheranostic nanoagents (PTNAs) were engineered by conjugating nanoparticles either with a photosensitizer (rose bengal or pyridyl porphyrin) or with an imaging agent (rhodamine B). The assembled PTNAs were revealed to possess good fluorescent properties and to generate singlet oxygen, and showed antimicrobial properties when irradiated with low-cost green LED light. All of the processes and properties (synthesis, antioxidant potential, percent conjugation, fluorescence, singlet oxygen generation, and antimicrobial photodynamic therapy) of PTNAs synthesized using <i>S. cumini</i> were characterized and compared with those synthesized using chitosan. The antimicrobial effect of photodynamic therapy using developed PTNAs was characterized by confocal laser scanning microscopy (CLSM). The development of bioinspired PTNAs described herein will find applications of photodynamic therapy for the treatment of cancer and microbial infection

Design and Synthesis of Metalloporphyrin Nanoconjugates for Dual Light-Responsive Antimicrobial Photodynamic Therapy

Antimicrobial photodynamic therapy (APDT) utilizes photosensitizers (PSs) that eradicate a broad spectrum of bacteria in the presence of light and molecular oxygen. On the other hand, some light sources such as ultraviolet (UVB and UVC) have poor penetration and high cytotoxicity, leading to undesired PDT of the PSs. Herein, we have synthesized conjugatable mesosubstituted porphyrins and extensively characterized them. Time-dependent density functional theory (TD-DFT) calculations revealed that metalloporphyrin EP (5) is a suitable candidate for further applications. Subsequently, the metalloporphyrin was conjugated with lignin-based zinc oxide nanocomposites (ZnOAL and ZnOKL) to develop hydrophilic nanoconjugates (ZnOAL@EP and ZnOKL@EP). Upon dual light (UV + green light) exposure, nanoconjugates showed enhanced singlet oxygen generation ability and also demonstrated pH responsiveness. These nanoconjugates displayed significantly improved APDT efficiency (4–7 fold increase) to treat bacterial infection under dual light irradiation

Development of Gelatin Nanoparticle-Based Biodegradable Phototheranostic Agents: Advanced System to Treat Infectious Diseases

Rose bengal (RB)-conjugated and -entrapped gelatin nanoparticle (GNP)-based biodegradable nanophototheranostic (Bd-NPT) agents have been developed for the efficient antimicrobial photodynamic therapy. The study reveals that the use of gelatin nanoparticles could bypass the chemicals such as potassium iodide, EDTA, calcium chloride and polymyxin nonapeptide for the penetration of drug into the cell membrane to achieve antimicrobial activity. We demonstrated that the singlet oxygen generated by the biodegradable gelatin nanoparticles (BdGNPs) could damage the microbial cell membrane and the cell dies. The key features of the successive development of this work include the environmentally benign amidation of RB with GNPs, which was so far unexplored, and the entrapment of RB into the gelatin nanoparticles (GNP). The RB-GNP exhibited potent and broad-spectrum antimicrobial activity and could be useful in treating multi-drug-resistant microbial infections

Extraction of rutin from tagetes erecta (Marigold) and preparation of peroral nano-suspension for effective antitussive/expectorant therapy

The present study narrates the extraction of rutin from Tagetes erecta (Marigold) via maceration followed by ultrasonication. The extracted rutin was further fabricated into nanoparticles by high-pressure homogenization (HPH) and assessed by HPLC, DSC, XRD, TEM, and FTIR spectroscopy. The optimized batch of nanoparticles obtained using 32 central composite design (CCD) which exhibited particle size 209±14 nm, PDI 0.234±0.06, and 92±1.3% entrapment efficiency. The lyophilized rutin nanoparticles were further converted into nano-suspension. Interestingly, the rutin nano-suspension exhibited a similar antitussive effect in vivo as that by standard treatment pentoxyverine and reduced the coughing times within 2 min. Also, the phlegm showed high UV absorbance, implying its better expectorant activity than the standard and control. The rutin nano-suspension was highly stable and shelf life was found to be ∼29.1 months. The present study, for the first time, paves a way for the use of rutin nano-suspension to overcome chest congestion, shortening of breath, and in the management of cough

Bioinspired Nanotheranostic Agents: Synthesis, Surface Functionalization, and Antioxidant Potential

Bioinspired synthesis of nanomaterials is highly advantageous as a natural and cost-effective resource. Development of noble metal nanotheranostic agents was achieved through bioinspired synthetic routes. These biosynthesized nanoparticles were characterized by various analytical techniques including absorption spectroscopy, FTIR and electron microscopy (SEM and TEM). A large number of medicinal plants were screened, among which <i>Potentilla fulgens</i> (PF, vajradanti) and <i>Camellia sinensis</i> (CS, green tea) were found to produce nanomaterials with higher yields. Plant (PF and CS) mediated metallic nanoparticles had added advantage of metal reduction and simultaneous phytochemical capping over chemically synthesized procedures, which require multiple reagents. Antioxidant potential of the nanomaterials was determined by in vitro antioxidant assays confirming substantial antioxidant properties, which was due to the presence of phytochemicals on the nanoparticle surface. Flavonoids and catechins on the nanomaterial surface served as the supplier of hydroxyl groups for further derivatization. The surface of the nanoparticles was engineered by conjugating imaging and therapeutic moieties, resulting in the formation of theranostic nanoagents. The multimodal agents were characterized and the extent of drug loading was determined to validate the efficacy of those nanoconjugates. These bioinspired multimodal nanoprobes can serve as essential diagnostic and therapeutic tools in ongoing biomedical research