Acute hypoxia induces apoptosis of pancreatic β-cell by activation of the unfolded protein response and upregulation of CHOP

The success of pancreatic β-cells transplantation to treat type 1 diabetes has been hindered by massive β-cell dysfunction and loss of β-cells that follows the procedure. Hypoxia-mediated cell death has been considered one of the main difficulties that must be overcome for transplantation to be regarded as a reliable therapy. Here we have investigated the mechanisms underlying β-cell death in response to hypoxia (1% O2). Our studies show that mouse insulinoma cell line 6 (Min6) cells undergo apoptosis with caspase-3 activation occurring as early as 2 h following exposure to hypoxia. Hypoxia induces endoplasmic reticulum stress in Min6 cells leading to activation of the three branches of the unfolded protein response pathway. In response to hypoxia the pro-apoptotic transcription factor C/EBP homologous protein (CHOP) is upregulated. The important role of CHOP in the apoptotic process was highlighted by the rescue of Min6 cells from hypoxia-mediated apoptosis observed in CHOP-knockdown cells. Culturing isolated pancreatic mouse islets at normoxia showed intracellular hypoxia with accumulation of hypoxia-inducible factor-1α and upregulation of CHOP, the latter one occurring as early as 4 h after isolation. Finally, we observed that pancreatic islets of type 2 db/db diabetic mice were more hypoxic than their counterpart in normoglycemic animals. This finding indicates that hypoxia-mediated apoptosis may occur in type 2 diabetes

Fluorescence imaging through dynamic scattering media with speckle-encoded ultrasound-modulated light correlation

Fluorescence imaging is indispensable to biomedical research, and yet it remains challenging to image through dynamic scattering samples. Techniques that combine ultrasound and light as exemplified by ultrasound-assisted wavefront shaping have enabled fluorescence imaging through scattering media. However, the translation of these techniques into in vivo applications has been hindered by the lack of high-speed solutions to counter the fast speckle decorrelation of dynamic tissue. Here, we report an ultrasound-enabled optical imaging method that instead leverages the dynamic nature to perform imaging. The method utilizes the correlation between the dynamic speckle-encoded fluorescence and ultrasound-modulated light signal that originate from the same location within a sample. We image fluorescent targets with an improved resolution of ≤75 µm (versus a resolution of 1.3 mm with direct optical imaging) within a scattering medium with 17 ms decorrelation time. This new imaging modality paves the way for fluorescence imaging in highly scattering tissue in vivo

Cigarette smoke induces endoplasmic reticulum stress and the unfolded protein response in normal and malignant human lung cells

<p>Abstract</p> <p>Background</p> <p>Although lung cancer is among the few malignancies for which we know the primary etiological agent (i.e., cigarette smoke), a precise understanding of the temporal sequence of events that drive tumor progression remains elusive. In addition to finding that cigarette smoke (CS) impacts the functioning of key pathways with significant roles in redox homeostasis, xenobiotic detoxification, cell cycle control, and endoplasmic reticulum (ER) functioning, our data highlighted a defensive role for the unfolded protein response (UPR) program. The UPR promotes cell survival by reducing the accumulation of aberrantly folded proteins through translation arrest, production of chaperone proteins, and increased degradation. Importance of the UPR in maintaining tissue health is evidenced by the fact that a chronic increase in defective protein structures plays a pathogenic role in diabetes, cardiovascular disease, Alzheimer's and Parkinson's syndromes, and cancer.</p> <p>Methods</p> <p>Gene and protein expression changes in CS exposed human cell cultures were monitored by high-density microarrays and Western blot analysis. Tissue arrays containing samples from 110 lung cancers were probed with antibodies to proteins of interest using immunohistochemistry.</p> <p>Results</p> <p>We show that: 1) CS induces ER stress and activates components of the UPR; 2) reactive species in CS that promote oxidative stress are primarily responsible for UPR activation; 3) CS exposure results in increased expression of several genes with significant roles in attenuating oxidative stress; and 4) several major UPR regulators are increased either in expression (i.e., BiP and eIF2α) or phosphorylation (i.e., phospho-eIF2α) in a majority of human lung cancers.</p> <p>Conclusion</p> <p>These data indicate that chronic ER stress and recruitment of one or more UPR effector arms upon exposure to CS may play a pivotal role in the etiology or progression of lung cancers, and that phospho-eIF2α and BiP may have diagnostic and/or therapeutic potential. Furthermore, we speculate that upregulation of UPR regulators (in particular BiP) may provide a pro-survival advantage by increasing resistance to cytotoxic stresses such as hypoxia and chemotherapeutic drugs, and that UPR induction is a potential mechanism that could be attenuated or reversed resulting in a more efficacious treatment strategy for lung cancer.</p

The use of molecular probes for the characterization of dispersions of functionalized silica nanoparticles

Butoxylated silica nanoparticles (BSN) were prepared by esterification of the silanol groups of fumed silica nanoparticles with butanol and characterized by (13)C and (29)Si NMR and thermogravimetry. The molecular probes benzophenone (BP) and safranine-T were used to investigate the BSN suspensions in water: acetonitrile. Laser flash-photolysis experiments at lambda(exc)=266nm performed with BSN suspended in acetonitrile:aqueous phosphate buffer supported previous results of our group obtained by time-resolved phosphorescence experiments and showed that only free and adsorbed excited triplet states of BP and diphenylketyl radicals contribute to the signals. The UV-vis spectroscopic and photophysical properties of safranine-T are strongly solvent-dependent. Thus, the analysis of the emission spectra and fluorescence lifetimes yielded information on the localization of this probe molecule in suspensions of BSN and of the bare silica nanoparticles. The values of the equilibrium constant for the adsorption of the ground-state safranine-T on the particles were found to be (9.2 +/- 0.8) x 10(4), (7.2 +/- 0.8) x 10(5), and (3.0 +/- 0.1) X 10(4) for the BSN in 1: 1 acetonitrile:water, SiO(2) in 1:1 acetonitrile:water, and SiO(2) in acetonitrile, respectively. (C) 2009 Elsevier B.V. All rights reserved.7315460Agencia Nacional de Promocion Cientifica y Tecnologica, (ANPCyT, Argentina). [14508]DAAD Alumni ProgramConsejo Nacional de investigaciones Cientificas y Tecnicas (CONICET, Argentina)Comision de Investigaciones Cientificas de la Provincia de Buenos Aires (CICPBA, Argentina)Agencia Nacional de Promocion Cientifica y Tecnologica, (ANPCyT, Argentina). [14508

Safranine-T Triplet-State Quenching by Modified Silica Nanoparticles

Modified silica nanoparticles (NPs) were obtained by esterification of the silanol groups of fumed silica nanoparticles with 4-hydroxymethyl-N,N-dimethylbenzenamine. These particles were characterized by Fourier transform infrared spectroscopy, (13)C and (29)Si nuclear magnetic resonance, thermogravimetry, X-ray photoelectron spectroscopy, transmission electron microscopy with energy-dispersive spectroscopy, and measurement of the specific surface area. Laser flash photolysis experiments at the excitation wavelength of 532 nm were performed with safranine-T and NP suspensions in acetonitrile. Comparative experiments of safranine-T with solutions of 4-hydroxymethyl-N,N-dimethylbenzenamine and two other related alcohols were performed. Evidence for the triplet safranine-T H-abstraction from the organic group of the particles is obtained. The electron transfer from the organic group to the triplet excited states of safranine-T also takes place, as shown by the absorption traces obtained at 50 mu s after the laser shot.115371812218130Agencia Nacional de Promocion Cientifica y Tecnologica, (ANPCyT, Argentina) [0686]Agencia Nacional de Promocion Cientifica y Tecnologica, (ANPCyT, Argentina) [0686