Identification and Localization of Gold Nanoparticles in Potassium Ion Pores: Implications for Kir Blockade.

INTRODUCTION: In our previous study, we found that negatively charged gold nanoparticles with spermidine have the potential of blocking inwardly rectifying potassium channels (Kir), both at the cellular and the tissue level. METHODS: For the purpose of the present study, we purified the cytoplasmic domain of the Kir 3.1 receptor from Escherichia coli. Using single particles with surface coating by transmission electron microscopy, we identified the gold nanoparticles at the cytoplasmic side of the human Kir channel. RESULTS: Energy-dispersive X-ray spectroscopy showed the presence of the gold deposits in the cytoplasmic domain of the Kir receptor. CONCLUSION: In conclusion, we could identify undecagold in the ion pore of the Kir3.1 channel in order to clarify its direct blocking effect in the Kir ion pore by undecagold.

Gold nanoparticle-spermidine complex blocks the inward rectifier potassium channel.

A previous study showed that negatively charged gold nanoparticles block ion pores by binding to the sulfur group of the cysteine loop of the ion channel when small molecules like amine lead the nanoparticles inside the ion pore. Cells were voltage clamped at -100 mV. Subsequently a bath application of 30 µM Ach produced a current followed by the extracellular application of 100 mM spermidine and 50 nM of nanoparticle complex. Peak amplitude was then recorded. The addition of Ach (30 uM) reversed the effect, and we recorded inhibition of the peak amplitude. We also recorded electrocardiogram (EKG) and the atria effective refractory period (AERP) after treatment with the complex in the atrium of a rabbit heart in a Langendorff apparatus. Upon external application of the complex, the Ach-activated current was blocked by 48.8% ± 3.1% with 82.7% ± 3.1% reversal. In recording the EKG and the AERP after the addition of the complex including 30 mM spermidine with 50 nM nanoparticles, the complete resolution of atrial fibrillation at 50 s and the elongation of AERP from 46 to 52 was observed, which unveils a new class 3 anti arrythmic agent using gold nanoparticles with spermidine. Negatively charged gold nanoparticles (0.8 nm) block ion pores after penetrating the cell membrane with spermidine, thus entering the cells with a polyamine transporter and acting at the intracellular face of the channel via binding to the sulfur group of the human inward rectifying potassium channel- I(KAch).

Ileo-pelvic anastomosis and augmentation cystoplasty for treatment of encrusted pyelitis in a transplanted kidney.

Infection stones are more likely to form after urinary diversion as the result of urinary stasis. To prevent urinary stasis due to encrusted pyelitis in a transplanted kidney, we describe an alternative a surgical treatment: ileo-pelvic anastomosis. In our patient with a transplanted kidney, the ileal conduit had previously been anastomosed end-to-side owing to renal tuberculosis with an atrophied bladder; the transplanted ureter was anastomosed to the ileum in the left lower abdomen with an ileal conduit on the opposite side. Routine check-up revealed hydronephrosis with infected pyelitis and ureteritis in the transplanted kidney. We performed ileo-pelvic end-to-end anastomosis to prevent urinary stasis by lengthening the ileal conduit and performed augmentation cystoplasty to support the atrophied bladder following tuberculosis. We suggest that this approach may be useful in similar cases.

Gold nanoparticle-choline complexes can block nicotinic acetylcholine receptors.

We identified a novel class of direct ion-channel blockers of ligand-gated ion channels called the gold nanoparticle-choline complex. Negatively charged gold nanoparticles (1.4 nm) block ion pores by binding to the sulfur group of the cysteine loop of nicotinic acetylcholine receptors (nAChRs), and currents evoked by acetylcholine (Ach) can break these bonds. The current evoked by ACh in nAChRs was blocked directly in ion pores by the gold nanoparticle- choline complex. In adrenal-gland perfusion studies, the complex also blocked nAChRs by diminishing catecholamine release by about 75%. An in vivo study showed muscle relaxation in rats after injection of the complex. These results will foster the application of gold nanoparticles as a direct ion-channel blocker.

Identification of Novel Metabolic Proteins Released by Insulin Signaling of the Rat Hypothalmus Using Liquid Chromatography-Mass Spectrometry (LC-MS).

OBJECTIVE: The brain is dependent on glucose as an energy source. Intricate homeostatic mechanisms have been implicated in maintaining the blood glucose concentration in the brain. The aim of this study is to find the way to identify the metabolic proteins regulating the glucose in rat hypothalamus. METHODS: In this study, we analysed the secretome from rat hypothalamus in vivo. We introduced 500 nM of insulin into the rat hypothalamus. The chromatographic patterns of the secretome were identified, after which Mass Spectrometry-Mass Spectrometry (MS-MS) analysis was performed. RESULTS: In Liquid Chromatography-Mass Spectrometry (LC-MS) analysis, 60 proteins were identified in the secretome. Among them, 8 novel proteins were unveiled and were associated with the energy metabolism of insulin signaling in mitochondria of rat hypothalamic neuron. Nineteen other proteins have unknown functions. These ligands were confirmed to be secreting from the rat hypothalmus on insulin signaling by western blotting. CONCLUSION: The hypothalamus is the master endocrine gland responsible for the regulation of various physiological and metabolic processes. Proteomics using LC-MS analysis offer a efficient means for generating a comprehensive analysis of hypothalamic protein expression by insulin signaling.

Radiosensitization by targeting radioresistance-related genes with protein kinase A inhibitor in radioresistant cancer cells.

Here we determined which radiation-responsive genes were altered in radioresistant CEM/IR and FM3A/IR variants, which showed higher resistance to irradiation than parental human leukemia CEM and mouse mammary carcinoma FM3A cells, respectively and studied if radioresistance observed after radiotherapy could be restored by inhibition of protein kinase A. The expressions of DNA-PKcs, Ku70/80, Rad51 and Rad54 genes that related to DNA damage repair, and Bcl-2 and NF-kappaB genes that related to antiapoptosis, were up-regulated, but the expression of proapototic Bax gene was down-regulated in the radioresistant cells as compared to each parental counterpart. We also revealed that the combined treatment of radiation and the inhibitor of protein kinase A (PKA) to these radioresistant cells resulted in synergistic inhibition of DNA-PK, Rad51 and Bcl-2 expressions of the cells, and consequently restored radiosensitivity of the cells. Our results propose that combined treatment with radiotherapy and PKA inhibitor can be a novel therapeutic strategy to radioresistant cancers.