Transmission Electron Microscopy of Lipid Vesicles for Drug Delivery: Comparison between Positive and Negative Staining

Bello, Valentina; Mattei, Giovanni; Mazzoldi, P.; Vivenza, Nicoletta; Gasco, Paolo; Idée, Jean Marc; Robic, Caroline; Borsella, E.

doi:10.1017/s1431927610093645

Cited by 33 publications

(21 citation statements)

References 25 publications

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“…With a two-step staining procedure with UAR followed by lead citrate (Reynolds, 1963) in TEM, we could visualize the development of a cuticular lipid layer in E. pusilla fruits. Cross sections of 3D reconstructions of micro-CT scans of E. pusilla fruits, stained with phosphotungstic acid (PTA), a negative staining agent for lipids (Melchior et al, 1980; Bello et al, 2010), also revealed a white-stained layer at the dehiscence zones of the valves of E. pusilla (Figure 7). On the other hand, in both terrestrial species a layer of lignified cells can be seen at the valve margins (Figures 5H,J).…”

Section: Discussionmentioning

confidence: 99%

Morphological and Molecular Characterization of Orchid Fruit Development

et al. 2019

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Efficient seed dispersal in flowering plants is enabled by the development of fruits, which can be either dehiscent or indehiscent. Dehiscent fruits open at maturity to shatter the seeds, while indehiscent fruits do not open and the seeds are dispersed in various ways. The diversity in fruit morphology and seed shattering mechanisms is enormous within the flowering plants. How these different fruit types develop and which molecular networks are driving fruit diversification is still largely unknown, despite progress in eudicot model species. The orchid family, known for its astonishing floral diversity, displays a huge variation in fruit dehiscence types, which have been poorly investigated. We undertook a combined approach to understand fruit morphology and dehiscence in different orchid species to get more insight into the molecular network that underlies orchid fruit development. We describe fruit development in detail for the epiphytic orchid species Erycina pusilla and compare it to two terrestrial orchid species: Cynorkis fastigiata and Epipactis helleborine . Our anatomical analysis provides further evidence for the split carpel model, which explains the presence of three fertile and three sterile valves in most orchid species. Interesting differences were observed in the lignification patterns of the dehiscence zones. While C. fastigiata and E. helleborine develop a lignified layer at the valve boundaries, E. pusilla fruits did not lignify at these boundaries, but formed a cuticle-like layer instead. We characterized orthologs of fruit-associated MADS-domain transcription factors and of the Arabidopsis dehiscence-related genes INDEHISCENT (IND)/HECATE 3 (HEC3), REPLUMLESS (RPL) and SPATULA (SPT)/ALCATRAZ (ALC) in E. pusilla , and found that the key players of the eudicot fruit regulatory network appear well-conserved in monocots. Protein-protein interaction studies revealed that MADS-domain complexes comprised of FRUITFULL (FUL), SEPALLATA (SEP) and AGAMOUS (AG) /SHATTERPROOF (SHP) orthologs can also be formed in E. pusilla , and that the expression of HEC3, RPL , and SPT can be associated with dehiscence zone development similar to Arabidopsis. Our expression analysis also indicates differences, however, which may underlie fruit divergence.

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Section: Discussionmentioning

confidence: 99%

Morphological and Molecular Characterization of Orchid Fruit Development

et al. 2019

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“…IEM was performed as described with a few modifications [17–18]. Briefly, cells were fixed with 4% paraformaldehyde and 1% glutaral in 0.1 M phosphate buffer at 4°C for 4 hrs.…”

Section: Methodsmentioning

confidence: 99%

Secretion of annexin A3 from ovarian cancer cells and its association with platinum resistance in ovarian cancer patients

Yin

Yan

Yao

et al. 2012

J Cellular Molecular Medi

107

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Early detection of resistance to platinum-based therapy is critical for improving the treatment of ovarian cancers. We have previously found that increased expression of annexin A3 is a mechanism for platinum resistance in ovarian cancer cells. Here we demonstrate that annexin A3 can be detected in the culture medium of ovarian cancer cells, particularly these cells that express high levels of annexin A3. Levels of annexin A3 were then determined in sera from ovarian cancer patients using an enzyme-linked immunosorbent assay. Compared with those from normal donors, sera from ovarian cancer patients contain significantly higher levels of annexin A3. Furthermore, serum levels of annexin A3 were significantly higher in platinum-resistant patients than in platinum-sensitive patients. To gain insight into the mechanism of secretion, the ovarian cancer cell lines were examined using both transmission electron microscopy and immunoelectron microscopy. Compared with parent cells, there are significantly more vesicles in the cytoplasm of ovarian cancer cells that express high levels of annexin A3, and at least some vesicles are annexin A3-positive. Moreover, some vesicles appear to be fused with the cell membrane, suggesting that annexin A3 secretion may be associated with exocytosis and the release of exosomes. This is supported by our observation that ovarian cancer cells expressing higher levels of annexin A3 released increased numbers of exosomes. Furthermore, annexin A3 can be detected in exosomes released from cisplatin-resistant cells (SKOV3/Cis) by immunoblotting and immunoelectron microscopy.

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“…The visibility in TEM of many lipid-based nanoconstructs may be efficiently improved by the application of osmium tetroxide ( Figure 1 c) [ 60 , 72 , 78 , 99 , 100 , 153 ], which induces the formation of an intensely electron-dense reduction product thanks to its addition to the double carbon-to-carbon bonds present in the lipid molecules [ 154 ].…”

Section: Ultrastructural Histochemistry In Nanomedical Researchmentioning

confidence: 99%

Transmission Electron Microscopy as a Powerful Tool to Investigate the Interaction of Nanoparticles with Subcellular Structures

Malatesta

2021

IJMS

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Nanomedical research necessarily involves the study of the interactions between nanoparticulates and the biological environment. Transmission electron microscopy has proven to be a powerful tool in providing information about nanoparticle uptake, biodistribution and relationships with cell and tissue components, thanks to its high resolution. This article aims to overview the transmission electron microscopy techniques used to explore the impact of nanoconstructs on biological systems, highlighting the functional value of ultrastructural morphology, histochemistry and microanalysis as well as their fundamental contribution to the advancement of nanomedicine.

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Transmission Electron Microscopy of Lipid Vesicles for Drug Delivery: Comparison between Positive and Negative Staining

Cited by 33 publications

References 25 publications

Morphological and Molecular Characterization of Orchid Fruit Development

Morphological and Molecular Characterization of Orchid Fruit Development

Secretion of annexin A3 from ovarian cancer cells and its association with platinum resistance in ovarian cancer patients

Transmission Electron Microscopy as a Powerful Tool to Investigate the Interaction of Nanoparticles with Subcellular Structures

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