| 1. |
- Bergstrand, Jan, et al.
(författare)
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Fast, streamlined fluorescence nanoscopy resolves rearrangements of SNARE and cargo proteins in platelets co-incubated with cancer cells
- 2022
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Ingår i: Journal of Nanobiotechnology. - : Springer Nature. - 1477-3155. ; 20:1
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Tidskriftsartikel (refereegranskat)abstract
- Background Increasing evidence suggests that platelets play a central role in cancer progression, with altered storage and selective release from platelets of specific tumor-promoting proteins as a major mechanism. Fluorescence-based super-resolution microscopy (SRM) can resolve nanoscale spatial distribution patterns of such proteins, and how they are altered in platelets upon different activations. Analysing such alterations by SRM thus represents a promising, minimally invasive strategy for platelet-based diagnosis and monitoring of cancer progression. However, broader applicability beyond specialized research labs will require objective, more automated imaging procedures. Moreover, for statistically significant analyses many SRM platelet images are needed, of several different platelet proteins. Such proteins, showing alterations in their distributions upon cancer progression additionally need to be identified. Results A fast, streamlined and objective procedure for SRM platelet image acquisition, analysis and classification was developed to overcome these limitations. By stimulated emission depletion SRM we imaged nanoscale patterns of six different platelet proteins; four different SNAREs (soluble N-ethylmaleimide factor attachment protein receptors) mediating protein secretion by membrane fusion of storage granules, and two angiogenesis regulating proteins, representing cargo proteins within these granules coupled to tumor progression. By a streamlined procedure, we recorded about 100 SRM images of platelets, for each of these six proteins, and for five different categories of platelets; incubated with cancer cells (MCF-7, MDA-MB-231, EFO-21), non-cancer cells (MCF-10A), or no cells at all. From these images, structural similarity and protein cluster parameters were determined, and probability functions of these parameters were generated for the different platelet categories. By comparing these probability functions between the categories, we could identify nanoscale alterations in the protein distributions, allowing us to classify the platelets into their correct categories, if they were co-incubated with cancer cells, non-cancer cells, or no cells at all. Conclusions The fast, streamlined and objective acquisition and analysis procedure established in this work confirms the role of SNAREs and angiogenesis-regulating proteins in platelet-mediated cancer progression, provides additional fundamental knowledge on the interplay between tumor cells and platelets, and represent an important step towards using tumor-platelet interactions and redistribution of nanoscale protein patterns in platelets as a basis for cancer diagnostics.
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| 2. |
- Bergstrand, Jan, et al.
(författare)
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Super-resolution microscopy can identify specific protein distribution patterns in platelets incubated with cancer cells
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Protein contents in platelets are frequently changed upon tumor development and metastasis. However, how cancer cells can influence protein-selective redistribution and release within platelets, thereby promoting tumor development, remains largely elusive. With fluorescence-based super-resolution stimulated emission depletion (STED) imaging we reveal how specific proteins, implicated in tumor progression and metastasis, re-distribute within platelets, when subject to soluble activators (thrombin, adenosine-diphosphate and thromboxaneA2), and when incubated with cancer (MCF-7, MDA-MB-231, EFO21) or non-cancer cells (184A1, MCF10A). Upon cancer cell incubation, the cell-adhesion protein P-selectin was found to re-distribute into circular nano-structures, consistent with accumulation into the membrane of protein-storing alpha-granules within the platelets. These changes were to a significantly lesser extent, if at all, found in platelets incubated with normal cells, or in platelets subject to soluble platelet activators. From these patterns, we developed a classification procedure, whereby platelets exposed to cancer cells, to non-cancer cells, soluble activators as well as non-activated platelets all could be identified in an automatic, objective manner. We demonstrate that STED imaging, in contrast to electron and confocal microscopy, has the necessary spatial resolution and labelling efficiency to identify protein distribution patterns in platelets and can resolve how they specifically change upon different activations. Combined with image analyses of specific protein distribution patterns within the platelets, STED imaging can thus have a role in future platelet-based cancer diagnostics and therapeutic monitoring. The presented approach can also bring further clarity into fundamental mechanisms for cancer cell-platelet interactions, and into non-contact cell-to-cell interactions in general.
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| 3. |
- Bergstrand, Jan, et al.
(författare)
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Super-resolution microscopy can identify specific protein distribution patterns in platelets incubated with cancer cells
- 2019
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Ingår i: Nanoscale. - : ROYAL SOC CHEMISTRY. - 2040-3364 .- 2040-3372. ; 11:20, s. 10023-10033
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Tidskriftsartikel (refereegranskat)abstract
- Protein contents in platelets are frequently changed upon tumor development and metastasis. However, how cancer cells can influence protein-selective redistribution and release within platelets, thereby promoting tumor development, remains largely elusive. With fluorescence-based super-resolution stimulated emission depletion (STED) imaging we reveal how specific proteins, implicated in tumor progression and metastasis, re-distribute within platelets, when subject to soluble activators (thrombin, adenosine diphosphate and thromboxane A2), and when incubated with cancer (MCF-7, MDA-MB-231, EFO21) or non-cancer cells (184A1, MCF10A). Upon cancer cell incubation, the cell-adhesion protein P-selectin was found to re-distribute into circular nano-structures, consistent with accumulation into the membrane of protein-storing alpha-granules within the platelets. These changes were to a significantly lesser extent, if at all, found in platelets incubated with normal cells, or in platelets subject to soluble platelet activators. From these patterns, we developed a classification procedure, whereby platelets exposed to cancer cells, to non-cancer cells, soluble activators, as well as non-activated platelets all could be identified in an automatic, objective manner. We demonstrate that STED imaging, in contrast to electron and confocal microscopy, has the necessary spatial resolution and labelling efficiency to identify protein distribution patterns in platelets and can resolve how they specifically change upon different activations. Combined with image analyses of specific protein distribution patterns within the platelets, STED imaging can thus have a role in future platelet-based cancer diagnostics and therapeutic monitoring. The presented approach can also bring further clarity into fundamental mechanisms for cancer cell-platelet interactions, and into non-contact cell-to-cell interactions in general.
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| 4. |
- Han, Hongya, et al.
(författare)
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Human 15-lipoxygenase-1 is a regulator of dendritic-cell spreading and podosome formation
- 2017
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Ingår i: The FASEB Journal. - : FEDERATION AMER SOC EXP BIOL. - 0892-6638 .- 1530-6860. ; 31:2, s. 491-504
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Tidskriftsartikel (refereegranskat)abstract
- Dendritic cells (DCs) involved in proinflammatory immune responses derive mainly from peripheral monocytes, and the cells subsequently mature and migrate into the inflammatory micromilieu. Here we report that suppressing of 15-lipoxygenase-1 led to a substantial reduction in DC spreading and podosome formation in vitro. The surface expression of CD83 was significantly lower in both sh-15-lipoxygenase-1 (15-LOX-1)-transduced cells and DCs cultivated in the presence of a novel specific 15-LOX-1 inhibitor. The T-cell response against tetanus-pulsed DCs was only affected to a minor extent on inhibition of 15-LOX-1. In contrast, endocytosis and migration ability of DCs were significantly suppressed on 15-LOX-1 inhibition. The expression of 15-LOX-1 in DCs was also demonstrated in affected human skin in atopic and contact dermatitis, showing that the enzyme is indeed expressed in inflammatory diseases in vivo. This study demonstrated that inhibiting 15-LOX-1 led to an impaired podosome formation in DCs, and consequently suppressed antigen uptake and migration capacity. These results indicated that 15-LOX-1 is a potential target for inhibiting the trafficking of DCs to lymphoid organs and inflamed tissues and decreasing the inflammatory response attenuating symptoms of certain immunologic and inflammatory disorders such as dermatitis.-Han, H., Liang, X., Ekberg, M., Kritikou, J. S., Brunnstro " m, angstrom., Pelcman, B., Matl, M., Miao, X., Andersson, M., Yuan, X., Schain, F., Parvin, S., Melin, E., Sjoberg, J., Xu, D., Westerberg, L. S., Bjorkholm, M., Claesson, H.- E. Human 15-lipoxygenase- 1 is a regulator of dendritic-cell spreading and podosome formation.
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| 5. |
- Miao, Xinyan
(författare)
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Platelet angiogenic activities : a concert of multiple players
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Angiogenesis, the sprouting of new capillaries from pre-existing blood vessels, is crucial for many physiological and pathological processes. Platelets are an important player in angiogenesis, whilst understanding of platelet engagement remains obscure. The present thesis aims to further elucidate the mechanisms of platelet angiogenic activities, including those by platelet releasate, platelet membrane, and miRNA-regulated platelet angiogenic factor synthesis. Our earlier studies have demonstrated that thrombin protease activated receptor 1 (PAR1) and PAR4 stimuli induce selective platelet release of proangiogenic and antiangiogenic regulators, respectively. The present thesis work shows that both PAR1-stimulated platelet releasate (PAR1-PR) and PAR4-PR enhanced tube formation and migration of endothelial progenitor cells (EPCs, also termed as endothelial colony forming cells, ECFCs, in paper II and IV) in vitro and angiogenesis in vivo, while the enhancements by PAR1-PR were more potent than those of PAR4-PR. Our findings suggest that the selective release of platelet angiogenic regulators may mainly concern the different release levels of platelet angiogenic regulators, and the final outcome of angiogenic regulating effects will depend on the negotiation of all factors in the platelet releasates. We further demonstrated that platelet-promoted angiogenesis depends on not only platelet-released mediators but also platelet membrane components. Thus, we found that platelet-enhanced EPC angiogenic responses were more pronounced than those by platelet releasates. We showed that platelet membrane glycoproteins had a major role in promoting EPC tube formation, and have identified that platelet tetraspanin CD151 and the integrin α6β1 expressed on both platelets and EPCs contributing to platelet-promoted tube formation. Albeit as anucleated cells, platelets retain certain capacities of protein synthesis. Our work gives further evidence that thrombin stimulation induced de novo synthesis of thrombospondin 1 (TSP1). More importantly, we showed that thrombin stimulation altered platelet miroRNA profile, and that thrombin-reduced platelet miR-27b expression enhanced TSP1 synthesis. The latter was supported by our data showing that miR-27b mimic transfection inhibited TSP1 synthesis in the platelet precursor megakaryocytes. We have also studied if type 2 diabetes mellitus (T2DM) would impair platelet angiogenic activities. We found that platelets from mild T2DM subjects were hyperreactive, but had similar angiogenic activities (angiogenic regulator release and platelet-regulated EPC tube formation), as compared to those of age/gender-matched controls. Our data suggest that a good glucose control may be beneficial for maintaining platelet angiogenic function. Together, this thesis work shows the platelets can regulate angiogenesis via different elements, and that platelet angiogenic activities are a concert of multiple players, namely released angiogenic factors, membrane components, as well de novo angiogenic factor synthesis and its regulation by miRNAs.
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