Fluorescence Dynamics in the Endoplasmic Reticulum of a Live Cell: Time‐Resolved Confocal Microscopy

Ghosh, Shirsendu; Nandi, Shyamapada; Ghosh, Catherine; Bhattacharyya, Kankan

doi:10.1002/cphc.201600425

Cited by 26 publications

(34 citation statements)

References 55 publications

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“…34,[45][46]49 It revealed that the intracellular biomembranes are significantly more polar (less ordered) than plasma membranes. 46 Second approach exploited specific targeting of solvatochromic dyes to organelle of interest, 5,25,[35][36][50][51][52][53] including mitochondria 32,50,54 and ER. 35 However, these studies were focused on targeting one particular organelle.…”

Section: Introductionmentioning

confidence: 99%

“…46 Second approach exploited specific targeting of solvatochromic dyes to organelle of interest, 5,25,[35][36][50][51][52][53] including mitochondria 32,50,54 and ER. 35 However, these studies were focused on targeting one particular organelle. Specific targeting of different organelles using a family of probes based on the same solvatochromic dye would enable direct comparison of properties of organelles and their response to different stress conditions.…”

Section: Introductionmentioning

confidence: 99%

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Targeted Solvatochromic Fluorescent Probes for Imaging Lipid Order in Organelles under Oxidative and Mechanical Stress

2021

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Biomembranes constitute a basis for all compartments of live cells, and therefore monitoring their lipid organization is essential for understanding cell status and activity. However, sensing and imaging lipid organization specifically in different organelles of live cells remains challenging. Here, we designed an array of solvatochromic probes based on Nile Red bearing ligands for specific targeting of endoplasmic reticulum, mitochondria, lysosomes, Golgi apparatus, plasma membranes and lipid droplets. These polaritysensitive probes detected variations in the lipid order by changing their emission maximum, as evidenced by fluorescence spectroscopy in model membranes. In colocalization microscopy experiments with reference organelle markers, they exhibited good organelle selectivity. Using two-color fluorescence microscopy, the new probes enabled imaging local polarity of organelles in live cells. To exclude the biased effect of the probe design on the sensitivity to membrane properties, we calibrated all probes in model membranes under the microscope, which enabled first quantitative description of lipid order in each organelle of interest. Cholesterol extraction/enrichment confirmed capacity of the probes to sense lipid order, revealing that organelles poor in cholesterol are particularly affected by its enrichment. The probes also revealed that oxidative and mechanical stresses produced changes in the local polarity and lipid order that were characteristic for each organelle, with mitochondria and lysosomes being particularly stresssensitive. The new probes constitute a powerful toolbox for monitoring the response of the cells to physical and chemical stimuli at the level of membranes of individual organelles, which remains an underexplored direction in the cellular research.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Targeted Solvatochromic Fluorescent Probes for Imaging Lipid Order in Organelles under Oxidative and Mechanical Stress

2021

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“…er provides a quality control system, regulating the modification and folding of membrane and secretory proteins and eliminating misfolded polypeptides through autophagic degradation or er-associated degradation (41). a variety of toxic insults including ca 2+ overload, failure of protein folding, synthesis, degradation or transport and hypoxia can interrupt the er function and give rise to erS (42).…”

Section: Discussionmentioning

confidence: 99%

Anticancer effect of caudatin in diethylnitrosamine‑induced hepatocarcinogenesis in rats

et al. 2020

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an overwhelming endoplasmic reticulum stress (erS) and the following unfolded protein response (uPr) can induce hepatic inflammation, fibrosis and hepatocellular carcinoma (Hcc). caudatin, one of the species of c-21 steroidal glycosides mainly isolated from the roots of Cynanchum bungei Decne, exhibits potent anticancer activities in vivo. However, the effect of caudatin on Hcc remains unclear. in the present study, a diethylnitrosamine (den)-induced Hcc model was established. Nodules and tumors in rat livers were monitored by T2-/T1-weighted-magnetic resonance imaging (MRI) using a 1.5 T scanner. Caudatin reduced the number and size of nodules and alleviated the inflammatory foci in the liver. In addition, the hepatic pro-inflammatory levels of interleukin (il) 6, monocyte chemoattractant protein 1 and il-1β were decreased in caudatin-treated rats. The den-induced surge in malondialdehyde, aspartate aminotransferase, alanine transaminase and TBil were alleviated following caudatin treatment. The expression of erS chaperones glucose-regulated protein, 94 kda, glucose-regulated protein, 78 kda and protein disulfide-isomerase A4 and the proliferation marker Ki-67 in liver nodules were all downregulated by caudatin as demonstrated by immunohistochemistry, reverse transcription-quantitative PCR and western blot analysis. Caudatin reduced the cytoprotective erS sensor activating transcription factor 6-mediated signal transduction and inhibited the PKr-like endoplasmic reticulum kinase/eukaryotic initiation factor 2α/activating transcription factor 4 pathway. However, the effect of caudatin on inositol requiring enzyme 1 signaling was negligible. In conclusion, restoration of the dysregulated uPr program was involved in the antitumor efficacy of caudatin without inducing cumulative hepatotoxicity.

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“…The HeLa cells were grown in phenol red free DMEM (Dulbecco's Modified Eagle Medium) with 10 % fetal bovine serum, 1 % Pen Strep Glutamine (from Gibco) in an atmosphere of 5 % (v/v) CO 2 enriched air at 37 °C, as described earlier elsewhere . For microscopy, a T25 flask have been utilized for the purpose of culture the cells.…”

Section: Methodsmentioning

confidence: 99%

Probing Viscosity of Co‐Polymer Hydrogel and HeLa Cell Using Fluorescent Gold Nanoclusters: Fluorescence Correlation Spectroscopy and Anisotropy Decay

et al. 2020

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Fluorescence dynamics of gold nanoclusters (Au9 and Au25) are studied in the complex and crowded environment of a triblock co‐polymer (F127) hydrogel and inside cervical cancer cell, HeLa. In the hydrogel, spherical micelles of F127 remain immobilized with a hydrophobic core (PPO) and a hydrophilic corona (PEO) region. The fluorescence anisotropy decay suggests that the timescale of rotational relaxation in the hydrogel is similar to that in bulk water (viscosity ∼1 cP). From fluorescence correlation spectroscopy (FCS) it is inferred that the local viscosity in the hydrogel is 12 cP for Au9 and 18 cP for Au23. These results indicate that gold nanoclusters (AuNCs) localize in the corona region of the hydrogel. Evidently, frictions against rotation and translation are different inside the gel. It is suggested that rotation of the AuNCs senses the immediate water‐like “void” region while translation motion involves in‐and‐out movement of the AuNCs at the periphery of the gel. Finally, the gold nanoclusters are used for cell imaging and estimation of intracellular viscosity of HeLa cells.

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Fluorescence Dynamics in the Endoplasmic Reticulum of a Live Cell: Time‐Resolved Confocal Microscopy

Cited by 26 publications

References 55 publications

Targeted Solvatochromic Fluorescent Probes for Imaging Lipid Order in Organelles under Oxidative and Mechanical Stress

Targeted Solvatochromic Fluorescent Probes for Imaging Lipid Order in Organelles under Oxidative and Mechanical Stress

Anticancer effect of caudatin in diethylnitrosamine‑induced hepatocarcinogenesis in rats

Probing Viscosity of Co‐Polymer Hydrogel and HeLa Cell Using Fluorescent Gold Nanoclusters: Fluorescence Correlation Spectroscopy and Anisotropy Decay

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