Raman microspectroscopy of live cells under autophagy-inducing conditions

Konorov, S. O.; Jardon, Mario A.; Piret, James M.; Blades, Michael W.; Turner, Robin F. B.

doi:10.1039/c2an35477b

Cited by 19 publications

(27 citation statements)

References 22 publications

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“…After apoptosis, necrosis or autophagy were induced by defined substances, we were able to identify unambiguously specific clusters for each type of cell death. The spectra of each group showed alterations in their peaks, which were highly specific for apoptosis, necrosis or autophagy . Morphological changes in the cells after treatment with the various drugs supported the induction of the relevant type of cell death (apoptosis, necrosis or autophagy).…”

Section: Discussionmentioning

confidence: 89%

“…The 3‐BrPv treatment group exhibited a decrease in all these peaks. In contrast to cells treated with staurosporine or 3‐BrPv, the resveratrol‐treated cells had less pronounced spectral changes; however, the spectra exhibited a clearly decreased signal at 718/cm (Table ), which is associated with phospholipids and was shown be a useful spectral marker of autophagic response …”

Section: Resultsmentioning

confidence: 99%

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Raman spectroscopy as an analytical tool for melanoma research

et al. 2014

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SummaryBackground. Raman spectroscopy is an optical noninvasive screening technology that generates individual fingerprints of living cells by reflecting their molecular constitution. Aim. To discriminate melanoma cells from melanocytes, to identify drug-induced melanoma cell death stages (apoptosis, necrosis, autophagy) and to assess the susceptibility of melanoma cells to anticancer therapy. Methods. We used Raman spectroscopy on normal and melanoma cells, and on wild-type (WT) and mutant melanoma cells, to investigate whether the technique could distinguish between different types of cells, identify mutations and evaluate response to anticancer therapy. Results. Using the multivariate principal component analysis of the Raman spectra, melanocytes could be distinguished from melanoma cells, and WT melanoma cells could be distinguished from melanoma cells with BRAF or NRAS mutations. When we used the apoptosis inducer staurosporine, the necrosis inducer 3-bromopyruvate and the autophagy inducer resveratrol to induce cell death in SKMEL28 melanoma cells, Raman spectroscopy clearly distinguished between these three types of cell death, as confirmed by immunoblotting. Finally, the technique could discriminate between different melanoma cell lines according to their susceptibility to high-dose ascorbate. Conclusions. Raman spectroscopy is a powerful noninvasive tool to distinguish between melanocytes and melanoma cells, to analyze the specific type of cell death in melanoma cells, and to predict the susceptibility of melanoma cells to anticancer drugs.

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Section: Discussionmentioning

confidence: 89%

Section: Resultsmentioning

confidence: 99%

Raman spectroscopy as an analytical tool for melanoma research

et al. 2014

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“…Using Raman spectroscopy, Konorov et al observed increased phospholipid levels in LMD cells undergoing autophagy due to glutamine depletion. These cells were found to accumulate lipids during autophagy.…”

Section: Resultsmentioning

confidence: 99%

Effects of cysteine, asparagine, or glutamine limitations in Chinese hamster ovary cell batch and fed‐batch cultures

Ghaffari

Jardon

Krahn

et al. 2019

Biotechnology Progress

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Amino acid availability is a key factor that can be controlled to optimize the productivity of fed‐batch cultures. To study amino acid limitation effects, a serum‐free chemically defined basal medium was formulated to exclude the amino acids that became depleted in batch culture. The effect of limiting glutamine, asparagine, and cysteine on the cell growth, metabolism, antibody productivity, and product glycosylation was investigated in three Chinese hamster ovary (CHO) cell lines (CHO‐DXB11, CHO‐K1SV, and CHO‐S). Cysteine limitation was detrimental to both cell proliferation and productivity for all three CHO cell lines. Glutamine limitation reduced growth but not cell specific productivity, whereas asparagine limitation had no significant effect on either growth or cell specific productivity. Neither glutamine nor asparagine limitation significantly affected antibody glycosylation. Replenishing the CHO‐DXB11 culture with cysteine after 1 day of cysteine limitation allowed the cells to partially recover their growth and productivity. This recovery was not observed after 2 days of cysteine limitation. Based on these findings, a fed‐batch protocol was developed using single or mixed amino acid supplementation. Although cell density and antibody concentration were lower compared to a commercial feed, the feeds based on cysteine supplementation yielded comparable cell specific productivity. Overall, this study showed that different amino acid limitations have varied effects on the performance of CHO cell cultures and that maintaining cysteine availability is a critical process parameter for the three cell lines investigated.

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“…Electron microscopy studies have indicated that complete deprivation of serum and amino acids provides a useful model for the further study of cellular autophagy [19] . It is widely accepted that autophagy is induced in several Autophagy induction Autophagy induction Autophagy induction Autophagy induction mTOR-dependent signaling pathway mTOR-dependent signaling pathway mTOR-dependent signaling pathway mTOR-dependent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway mTOR-dependent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway mTOR-independent signaling pathway [20,21] . Drosophila Rack1, a receptor of activated protein kinase C 1, increases 4.1-to 5.5-fold during nutrient deprivation in all three genotypes.…”

Section: Starvation Inducersmentioning

confidence: 99%

Application and interpretation of current autophagy inhibitors and activators

et al. 2013

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Autophagy is the major intracellular degradation system, by which cytoplasmic materials are delivered to and degraded in the lysosome. As a quality control mechanism for cytoplasmic proteins and organelles, autophagy plays important roles in a variety of human diseases, including neurodegenerative diseases, cancer, cardiovascular disease, diabetes and infectious and inflammatory diseases. The discovery of ATG genes and the dissection of the signaling pathways involved in regulating autophagy have greatly enriched our knowledge on the occurrence and development of this lysosomal degradation pathway. In addition to its role in degradation, autophagy may also promote a type of programmed cell death that is different from apoptosis, termed type II programmed cell death. Owing to the dual roles of autophagy in cell death and the specificity of diseases, the exact mechanisms of autophagy in various diseases require more investigation. The application of autophagy inhibitors and activators will help us understand the regulation of autophagy in human diseases, and provide insight into the use of autophagy-targeted drugs. In this review, we summarize the latest research on autophagy inhibitors and activators and discuss the possibility of their application in human disease therapy.

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Raman microspectroscopy of live cells under autophagy-inducing conditions

Cited by 19 publications

References 22 publications

Raman spectroscopy as an analytical tool for melanoma research

Raman spectroscopy as an analytical tool for melanoma research

Effects of cysteine, asparagine, or glutamine limitations in Chinese hamster ovary cell batch and fed‐batch cultures

Application and interpretation of current autophagy inhibitors and activators

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