Patch-Clamp Study of Single Ryanodine Receptor Channels in the Outer Nuclear Membrane

Poster presentationModulation of cytoplasmic free calcium (Ca2+) concentration is a universal signaling pathway that regulates numerous cellular processes. Ubiquitous intracellular Ca2+ release channels – inositol 1,4,5-trisphosphate receptor (InsP3R) and ryanodine receptor (RyR) channels – localized in the sarco/endoplasmic reticulum (ER) play a central role in this pathway in all animal cells. Electrophysiological study of the single-channel conductance and gating properties of these Ca2+ release channels with conventional patch-clamp approach has been hindered by their intracellular localization. To overcome this limitation, patch-clamp electrophysiology has been applied on isolated nuclei where these Ca2+ release channels are found abundantly in the outer nuclear envelope. We have successfully uterlized this nuclear membrane electrophysiology to study the gating properties of single InsP3R channels in several cellular systems. Whereas, all the current single channel data, including channel conductance, permeation properties, and ligand regulation, of the RyR channels were done exclusively by reconstituting the channels into artificial planar lipid bilayers. To gain insights into the single channel properties of the RyR in its native membrane milieu, we applied nuclear membrane electrophysiological study on isolated nuclei from stable-inducible mouse RyR2 HEK-293 cells. Using potassium as charge carrier, caffeine activated single channel current with conductance of 750 pS in isolated nuclei. This caffeine activated current showed a linear current/voltage relationship under symmetrical ionic conditions and was sensitive to non-specific RyR inhibitor, ruthenium red. Furthermore, the single RyR channels recorded from the outer nuclear membrane exhibited bi-phasic Ca2+ regulation. In conclusion, we demonstrated, for the first time, that single RyR channels recordings from isolated nuclei and our results suggested that the nuclear membrane electrophysiology could be a sensitive and robust technique to study the gating properties of intracellular channels, including the InsP3R and RyR.published_or_final_versio

Molecular Mechanism of Capacitative Calcium Entry Deficits in Familial Alzheimer’s Disease

Poster PresentationPresenilin (PS) is the catalytic subunit of the gamma-secretase which is responsible for the cleavage of amyloid precursor protein to form beta amyloid (Aβ). Mutations in PS associated with familial Alzheimer’s disease (FAD) increase the Aβ plaques formation in the brain and cause neurodegeneration. Apart from this, FAD-linked PS mutations have been demonstrated to disrupt intracellular calcium (Ca2+) regulation. Accumulating evidence suggests that Ca2+ disruption may play a proximal role in the AD pathogenesis. Mutant PS exaggerated Ca2+ release from the endoplasmic reticulum (ER). It also attenuated Ca2+ entry through the capacitative Ca2+ entry (CCE) pathway, yet, the mechanism is not fully understood. Using a human neuroblast cell line SH-SY5Y and Ca2+ imaging technique, we observed CCE deficits in FAD-linked PS1-M146L retroviral infected cell. The attenuation of CCE in PS1 mutant cells was not mediated by the down-regulation of STIM1 and Orai1 expression, the known essential molecular players in the CCE pathway. Instead, we identified a molecular interaction between PS and STIM1 proteins by immunoprecipitation. On the other hand, immunofluorescence staining showed a significant reduction in puncta formation after ER Ca2+ depleted by thapsigargin in cells infected with PS1-M146L as compared to the wild type PS1 infected cells. Taken together, our results suggest a molecular mechanism for the CCE deficits in FAD associated with PS1 mutations. The interaction of mutant PS1 with STIM1 exerts a negative impact on its oligomerization and/or its interaction with Orai1. Our results may suggest molecular targets for the development of therapeutic agents that help to treat the disease.published_or_final_versio

Is customer participation in value creation a double-edged sword? Evidence from professional financial services across cultures

2009-2010 > Academic research: refereed > Publication in refereed journalAccepted ManuscriptPublishe

Strengthening customer loyalty through intimacy and passion : roles of customer–firm affection and customer–staff relationships in services

2008-2009 > Academic research: refereed > Publication in refereed journalAccepted ManuscriptPublishe

Do customers and employees enjoy service participation? Synergistic effects of seif- and other-efficacy

2012-2013 > Academic research: refereed > Publication in refereed journalAccepted ManuscriptPublishe

Effect of an edible nanomultilayer coating by electrostatic self-assembly on the shelf life of fresh-cut mangoes

This work aims at evaluating the effect of an alginate-chitosan nanomultilayer coating, obtained by electrostatic layer-by-layer self-assembling, in the quality and shelf life of fresh-cut mangoes. Coated and uncoated fresh-cut mangoes were stored under refrigeration (8 °C) for 14 days. The changes in mass loss, titratable acidity, pH, ascorbic acid content, total soluble solids, malondialdehyde content, browning rate, and microbial count were evaluated during storage. At the end of the storage period, lower values of mass loss, pH, malondialdehyde content, browning rate, soluble solids, microorganisms proliferation, and higher titratable acidity were observed in the coated mangoes. The nanomultilayer coating did not improve the retention of vitamin C during storage of fresh-cut mangoes. Results suggest that chitosan-alginate nanomultilayer edible coating extends the shelf life of fresh-cut mangoes up to 8 days.Author Marthyna Pessoa de Souza thanks Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES/PDEE-Brazil) and Fundacao de Amparo a Ciencia e Tecnologia do Estado de Pernambuco (FACEPE, Brazil) for granting her scholarships. The authors thank the Fundacao para a Ciencia e a Tecnologia (FCT) Strategic Project PEst-OE/EQB/LA0023/2013 and the Project "BioInd-Biotechnology and Bioengineering for improved Industrial and Agro-Food processes", REF. NORTE-07-0124-FEDER-000028, co-funded by the Programa Operacional Regional do Norte (ON.2-O Novo Norte), QREN, and FEDER (Portugal)

Sleep behavior and depression: Findings from the China Kadoorie Biobank of 0.5 million Chinese adults.

BACKGROUND: Mixed results have shown the association between sleep behavior and depression, but evidence relating the joint effect of sleep duration and sleep disturbances is limited, especially in Chinese population. METHODS: A total of 512,891 adults aged 30-79 years from China Kadoorie Biobank (CKB) were included. Depression was defined by Composite International Diagnostic Inventory-short form (CIDI-SF). Sleep duration and sleep disturbances, including difficulty initiating and maintaining sleep (DIMS), early morning awakening (EMA), daytime dysfunction (DDF) and any sleep disturbances (ASD), were obtained by a self-reported questionnaire. Logistic regression was applied to examine the association between sleep behavior and depression. RESULTS: About 23.1% of participants reported short sleep duration (≤ 6h), and 5.1% reported long sleep duration (> 9h). Compared with normal sleep duration (7-9h), both groups were associated greater likelihood of having depression (short sleep: OR = 2.32, 95%CI: 2.14-2.51; long sleep: OR = 1.56, 96%CI: 1.34-1.81). Participants reported sleep disturbances were significantly associated with depression (odds ratios ranged from 3.31 to 4.17). Moreover, the associations tended to be stronger for those who reported both abnormal sleep duration and sleep disturbances (p for interactions < 0.05), especially for those who slept long. LIMITATIONS: The cross-sectional nature of the study design limits the interpretation of the results. CONCLUSIONS: Abnormal sleep duration and sleep disturbances were associated with depression. The associations were stronger for abnormal sleep duration accompanied with sleep disturbances, especially for a long duration. More attention should be paid on these persons in clinical practice

Pets as Sentinels of Human Exposure to Neurotoxic Metals

The idea that animals may be used as sentinels of environmental hazards pending over humans and the associated public health implications is not a new one. Nowadays pets are being used as bioindicators for the effects of environmental contaminants in human populations. This is of paramount importance due to the large increase in the worldwide distribution of synthetic chemicals, particularly in the built environment. Companion animals share the habitat with humans being simultaneously exposed to and suffering the same disease spectrum as their masters. Moreover, their shorter latency periods (due to briefer lifespans) enable them to act as early warning systems, allowing timely public health interventions. The rise on ethical constraints on the use of animals and, consequently, on the sampling they can be subjected to has led to the preferential use of noninvasive matrices, and in this case we are looking into hair. This chapter focuses in three non-essential metals: mercury, lead, and cadmium, due to their ubiquitous presence in the built environment and their ability of affecting the mammal nervous system. There is a fairly short amount of studies reporting the concentrations of these metals in pets’ hair, particularly for cats. These studies are characterized, and the metal concentrations corresponding to different parameters (e.g., age, sex, diet, rearing) are described in order to provide the reader with a general vision on the use of this noninvasive matrix on the studies conducted since the last two decades of the twentieth century.publishe