Single drop analysis of mercury ions by rational design of peptide coated gold nanoparticles integrated with MALDI-MS measurement.

In this study, a novel and rapid method for specific identification and accurate quantification of Hg2+ in environmental water was developed by using laser cleavable cysteine containing peptides modified gold nanoparticles coupled with high resolution matrix-assisted laser desorption ionization time-of-flight mass spectrometer (MALDI-TOF MS) measurement. First, gold nanoparticles were prepared by the reduction of tetrachloroauric (III) acid (HAuCl4) solution. Various cysteine containing peptides, photolabile linkers, including mercury ion binding motif with a proper molecular mass and amino acids were synthesized by solid phase peptide synthesis (SPPS). Subsequently, thiol-containing peptides were coated onto the surface of gold nanoparticles via the formation of gold-thiol (Au-S) bond. The resulting cysteine containing peptides modified gold nanoparticles were designed to specifically capture Hg2+ in water samples. After conjugated complex formation, ions of Hg2+-peptide complex were directly liberated by ultraviolet laser radiation by way of MALDI-MS using α-Cyano-4-hydroxycinnamic acid (CHCA) as matrix. The linear dynamic range of Hg2+ concentration in this study was 1-100 pmol/µL with coefficient of determination 0.9987. The limit of detection (LOD) and limit of quantification (LOQ) were 0.19 and 0.63 pmol/µL, respectively. Notably, the developed method allows rapid quantification of Hg2+ in 5 min and the desired sample volume was down to few µL.

Loss of PTEN causes SHP2 activation, making lung cancer cells unresponsive to IFN-γ.

Src homology-2 domain-containing phosphatase (SHP) 2, an oncogenic phosphatase, inhibits type II immune interferon (IFN)-γ signaling by subverting signal transducers and activators of transcription 1 tyrosine phosphorylation and activation. For cancer immunoediting, this study aimed to investigate the decrease of phosphatase and tensin homolog deleted on chromosome 10 (PTEN), a tumor suppressor protein, leading to cellular impairment of IFN-γ signaling. In comparison with human lung adenocarcinoma A549 cells, the natural PTEN loss in another human lung adenocarcinoma line, PC14PE6/AS2 cells, presents reduced responsiveness in IFN-γ-induced IFN regulatory factor 1 activation and CD54 expression. Artificially silencing PTEN expression in A549 cells also caused cells to be unresponsive to IFN-γ without affecting IFN-γ receptor expression. IFN-γ-induced inhibition of cell proliferation and cytotoxicity were demonstrated in A549 cells but were defective in PC14PE6/AS2 cells and in PTEN-deficient A549 cells. Aberrant activation of SHP2 by ROS was specifically shown in PC14PE6/AS2 cells and PTEN-deficient A549 cells. Inhibiting ROS and SHP2 rescued cellular responses to IFN-γ-induced cytotoxicity and inhibition of cell proliferation in PC14PE6/AS2 cells. These results demonstrate that a decrease in PTEN facilitates ROS/SHP2 signaling, causing lung cancer cells to become unresponsive to IFN-γ.

Autophagy facilitates cytokine-induced ICAM-1 expression.

ICAM-1 can be induced by inflammatory cytokines such as IFN-γ and TNF-α. This study investigated whether autophagy regulates ICAM-1 given that autophagy facilitates signaling of these two cytokines. Exogenous IFN-γ induced ICAM-1 in human lung epithelial A549 cells carrying wild type p53, a transcription factor reported for ICAM-1, but not in PC14PE6/AS2 (AS2) cells carrying mutated p53. However, IFN-γ also induced ICAM-1 in A549 cells with short hairpin RNA-silenced p53. No changes in IFN-γ receptor expression were observed in AS2 cells, but IFN-γ-activated Jak2/STAT1/IFN regulatory factor 1 was markedly decreased. In AS2 cells, increased levels of reactive oxygen species induced the activation of Src homology domain-containing phosphatase 2 (SHP2), while SHP2 was essential for IFN-γ resistance. AS2 cells showed autophagy resistance, and the manipulation of the autophagy pathway altered IFN-γ resistance. Aberrant Bcl-2 expression and mammalian target of rapamycin activation contributed to both autophagy resistance and IFN-γ resistance. Autophagy, but not p53, also modulated TNF-α-induced NF-κB activation and ICAM-1 expression. Inhibiting autophagy decreased the adhesion of human monocytic U937 cells to IFN-γ-treated A549 cells. These results demonstrated that IFN-γ and TNF-α induced ICAM-1 expression through a common pathway that was regulated by autophagy, but not p53.