RESUMEN
As the hearts of tumor cells, the nucleus is the ultimate target of many chemotherapeutic agents and genes. However, nuclear drug delivery is always hampered by multiple intracellular obstacles, such as low efficiency of lysosome escape and insufficient nuclear trafficking. Herein, an N-(2-hydroxypropyl) methacrylamide (HPMA) polymer-based drug delivery system was designed, which could achieve direct cytoplasmic delivery by a nonendocytic pathway and transport into the nucleus in a microtubules dependent fashion. A special targeting peptide (MT), derived from an endogenic parathyroid hormone-related protein, was conjugated to the polymer backbone, which could accumulate into the nucleus a by microtubule-mediated pathway. The in vitro studies found that low temperature and NaN3 could not influence the cell internalization of the conjugates. Besides, no obvious overlay of the conjugates with lysosome demonstrated that the polymer conjugates could enter the tumor cell cytoplasm by a nonendocytic pathway, thus avoiding the drug degradation in the lysosome. Furthermore, after suppression of the microtubule dynamics with microtubule stabilizing docetaxel (DTX) and destabilizing nocodazole (Noc), the nuclear accumulation of polymeric conjugates was significantly inhibited. Living cells fluorescence recovery after photobleaching study found that the nuclear import rate of conjugates was 2-fold faster compared with the DTX and Noc treated groups. These results demonstrated that the conjugates transported into the nucleus in a microtubules dependent way. Therefore, in addition to direct cytoplasmic delivery, our peptide conjugated polymeric platform could simultaneously mediate nuclear drug accumulation, which may open a new path for further intracellular genes/peptides delivery.
Asunto(s)
Citoplasma/metabolismo , Metacrilatos/química , Microtúbulos/metabolismo , Polímeros/química , Transporte Activo de Núcleo Celular/fisiología , Apoptosis/fisiología , Recuperación de Fluorescencia tras Fotoblanqueo , Células HeLa , Humanos , Microtúbulos/químicaRESUMEN
To improve the phosphorus (P) removal/recovery efficiency of a biological P removal system, the study used an alternating anaerobic/aerobic biofilter system to treat synthetic domestic sewage with a low carbon (C) to P ratio. The effects of using periodical carbon source amplification for P recovery on the P removal efficiency and the microbial characteristics within the biofilm were investigated. Intracellular storage polymer dyeing, scanning electron microscopy (SEM) observation and fluorescence in situ hybridization (FISH) methods were employed to characterize the changes of microbial communities in the biofilm during three continuous operation cycles of P bio-accumulating-P recovery (PB-PR). The results showed: through three cycles of operation process of PB-PR, the P removal efficiency of biofilter was increased from 60.3%, 82.9%, 86.6% (before P harvesting) to 87.2%, 91.2%, 93.5% (after P harvesting), respectively; the dominant microbial community morphotypes within the biofilter transformed from big cocci to small cocci, bacilli and filamentous and the group of phosphate-accumulating organisms (PAOs) got rising predominance, which was increased from 43% to 70% after three times of PB-PR; the proportion of PAOs in the biofilm increased unceasingly with the height of the up-flow biofilter. The results showed that the periodical carbon source amplification could improve the P removal efficiency of the biofilter and help the PAOs to become the dominant bacteria within the biofilm.