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Publications (10 of 14) Show all publications
Ahmed, M., Cerroni, B., Razuvaev, A., Härmark, J., Paradossi, G., Caidahl, K. & Gustafsson, B. (2017). Cellular Uptake of Plain and SPION-Modified Microbubbles for Potential Use in Molecular Imaging. Cellular and Molecular Bioengineering, 10(6), 537-548
Open this publication in new window or tab >>Cellular Uptake of Plain and SPION-Modified Microbubbles for Potential Use in Molecular Imaging
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2017 (English)In: Cellular and Molecular Bioengineering, ISSN 1865-5025, E-ISSN 1865-5033, Vol. 10, no 6, p. 537-548Article in journal (Refereed) Published
Abstract [en]

Both diagnostic ultrasound (US) and magnetic resonance imaging (MRI) accuracy can be improved by using contrast enhancement. For US gas-filled microbubbles (MBs) or silica nanoparticles (SiNPs), and for MRI superparamagnetic or paramagnetic agents, contribute to this. However, interactions of MBs with the vascular wall and cells are not fully known for all contrast media. We studied the in vitro interactions between three types of non-targeted air-filled MBs with a polyvinyl-alcohol shell and murine macrophages or endothelial cells. The three MB types were plain MBs and two types that were labelled (internally and externally) with superparamagnetic iron oxide nanoparticles (SPIONs) for US/MRI bimodality. Cells were incubated with MBs and imaged by microscopy to evaluate uptake and adhesion. Interactions were quantified and the MB internalization was confirmed by fluorescence quenching of non-internalized MBs. Macrophages internalized each MB type within different time frames: plain MBs 6 h, externally labelled MBs 25 min and internally labelled MBs 2 h. An average of 0.14 externally labelled MBs per cell were internalized after 30 min and 1.34 after 2 h; which was 113% more MBs than the number of internalized internally labelled MBs. The macrophages engulfed these three differently modified new MBs at various rate, whereas endothelial cells did not engulf MBs. Polyvinyl-alcohol MBs are not taken up by endothelial cells. The MB uptake by macrophages is promoted by SPION labelling, in particular external such, which may be important for macrophage targeting.

Place, publisher, year, edition, pages
SPRINGER, 2017
Keywords
Polyvinyl-alcohol, Macrophages, Endothelial cells, Interactions, In vitro
National Category
Medical Biotechnology
Identifiers
urn:nbn:se:kth:diva-217726 (URN)10.1007/s12195-017-0504-9 (DOI)000414198100003 ()2-s2.0-85027153168 (Scopus ID)
Note

QC 20171122

Available from: 2017-11-22 Created: 2017-11-22 Last updated: 2017-11-22Bibliographically approved
Härmark, J., Hebert, H. & Koeck, P. J. (2016). Shell thickness determination of polymer-shelled microbubbles using transmission electron microscopy. Micron, 85, 39-43
Open this publication in new window or tab >>Shell thickness determination of polymer-shelled microbubbles using transmission electron microscopy
2016 (English)In: Micron, ISSN 0968-4328, E-ISSN 1878-4291, Vol. 85, p. 39-43Article in journal (Refereed) Published
Abstract [en]

Intravenously injected microbubbles (MBs) can be utilized as ultrasound contrast agent (CA) resulting in enhanced image quality. A novel CA, consisting of air filled MBs stabilized with a shell of polyvinyl alcohol (PVA) has been developed. These spherical MBs have been decorated with superparamagnetic iron oxide nanoparticles (SPIONs) in order to serve as both ultrasound and magnetic resonance imaging (MRI) CA. In this study, a mathematical model was introduced that determined the shell thickness of two types of SPIONs decorated MBs (Type A and Type B). The shell thickness of MBs is important to determine, as it affects the acoustical properties. In order to investigate the shell thickness, thin sections of plastic embedded MBs were prepared and imaged using transmission electron microscopy (TEM). However, the sections were cut at random distances from the MB center, which affected the observed shell thickness. Hence, the model determined the average shell thickness of the MBs from corrected mean values of the outer and inner radii observed in the TEM sections. The model was validated using simulated slices of MBs with known shell thickness and radius. The average shell thickness of Type A and Type B MBs were 651nm and 637nm, respectively.

Place, publisher, year, edition, pages
Elsevier, 2016
Keywords
Shell thickness, Contrast agent, Transmission electron microscopy
National Category
Medical Engineering
Research subject
Medical Technology
Identifiers
urn:nbn:se:kth:diva-186876 (URN)10.1016/j.micron.2016.03.009 (DOI)000376217400006 ()27077316 (PubMedID)2-s2.0-84962835270 (Scopus ID)
Funder
EU, FP7, Seventh Framework Programme, 245572
Note

QC 20160516

Available from: 2016-05-16 Created: 2016-05-16 Last updated: 2017-11-30Bibliographically approved
Härmark, J. (2016). Structural studies of microbubbles and molecular chaperones using transmission electron microscopy. (Doctoral dissertation). Stockholm: KTH Royal Institute of Technology
Open this publication in new window or tab >>Structural studies of microbubbles and molecular chaperones using transmission electron microscopy
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Ultrasound contrast agents (CAs) are typically used in clinic for perfusion studies (blood flow through a specific region) and border delineating (differentiate borders between tissue structures) during cardiac imaging. The CAs used during ultrasound imaging usually consist of gas filled microbubbles (MBs) (diameter 1-5 μm) that are injected intravenously into the circulatory system. This thesis partially involves a novel polymer-shelled ultrasound CA that consists of air filled MBs stabilized by a polyvinyl alcohol (PVA) shell. These MBs could be coupled with superparamagnetic iron oxide nanoparticles (SPIONs) in order to serve as a combined CA for ultrasound and magnetic resonance imaging. The first three papers (Paper A-C) in this thesis investigate the structural characteristic and the elimination process of the CA.

In Paper A, two types (PVA Type A and PVA Type B) of the novel CA were analyzed using transmission electron microscopy (TEM) images of thin sectioned MBs. The images demonstrated that the SPIONs were either attached to the PVA shell surface (PVA Type A) or embedded in the shell (PVA Type B). The average shell thickness of the MBs was determined in Paper B by introducing a model that calculated the shell thickness from TEM images of cross-sectioned MBs. The shell thickness of PVA Type A was determined to 651 nm, whereas the shell thickness of PVA Type B was calculated to 637 nm. In Paper C, a prolonged blood elimination time was obtained for PVA-shelled MBs compared to the lipid-shelled CA SonoVue used in clinic. In addition, TEM analyzed tissue sections showed that the PVA-shelled MBs were recognized by the macrophage system. However, structurally intact MBs were still found in the circulation 24 h post injection. These studies illustrate that the PVA-shelled MBs are stable and offer large chemical variability, which make them suitable as CA for multimodal imaging.

This thesis also involves studies (Paper D-E) of the molecular chaperones (Hsp21 and DNAJB6). The small heat shock protein Hsp21 effectively protects other proteins from unfolding and aggregation during stress. This chaperone ability requires oligomerization of the protein. In Paper D, cryo-electron microscopy together with complementary structural methods, obtained a structure model which showed that the Hsp21 dodecamer (12-mer) is kept together by paired C-terminal interactions.The human protein DNAJB6 functions as a very efficient suppressor of polyglutamine (polyQ) and amyloid-β42 (Aβ42) aggregation. Aggregation of these peptides are associated with development of Huntington’s (polyQ) and Alzheimer’s (Aβ42) disease. In Paper E, a reconstructed map of this highly dynamic protein is presented, showing an oligomer with two-fold symmetry, indicating that the oligomers are assembled by two subunits.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2016. p. xiii, 58 s
Series
TRITA-STH : report, ISSN 1653-3836 ; 2016:3
Keywords
Transmission electron microscopy, Contrast agent, Microbubble, Polyvinyl alcohol, Single particle analysis, Heat shock protein, Molecular chaperone
National Category
Structural Biology
Research subject
Technology and Health
Identifiers
urn:nbn:se:kth:diva-186882 (URN)978-91-7729-020-9 (ISBN)
Public defence
2016-06-08, Hörsalen plan 4 Novum, Blickagången 6, 141 57, Huddinge, 10:00 (English)
Opponent
Supervisors
Note

QC 20160527

Available from: 2016-05-27 Created: 2016-05-16 Last updated: 2016-05-27Bibliographically approved
Härmark, J., Larsson, M. K., Razuvajev, A., Koeck, P. J., Paradossi, G., Brodin, L.-Å., . . . Bjällmark, A. (2015). Investigation of the elimination process of a multimodal polymer-shelled contrast agent in rats using ultrasound and transmission electron microscopy. Biomedical Spectroscopy and Imaging, 4(1), 81-93
Open this publication in new window or tab >>Investigation of the elimination process of a multimodal polymer-shelled contrast agent in rats using ultrasound and transmission electron microscopy
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2015 (English)In: Biomedical Spectroscopy and Imaging, ISSN 2212-8794, Vol. 4, no 1, p. 81-93Article in journal (Refereed) Published
Abstract [en]

BACKGROUND: A novel polymer-shelled contrast agent (CA) with multimodal imaging and target specific potential was developed recently and tested for its acoustical properties using different in-vitro setups.

OBJECTIVE: The aim of this study was to investigate the elimination of three types of the novel polymer-shelled CA, one unmodified and two shell modified versions, in rats.

METHODS: The blood elimination time was estimated by measuring the image intensity, from ultrasound images of the common carotid artery, over time after a bolus injection of the three types of the novel CA. The commercially available CA SonoVue was used as a reference. The subcellular localization of the three CAs was investigated using transmission electron microscopy.

RESULTS: The ultrasound measurements indicated a blood half-life of 17–85 s for the different types of the novel CA, which was significant longer than the blood half-life time for SonoVue. Additionally, CAs were exclusively found in the circulatory system, either taken up by, or found in the vicinity of macrophages.

CONCLUSIONS: Compared to the commercially available CA SonoVue, the blood circulation times for the three types of the novel polymer-shelled CA were prolonged. Moreover, macrophages were suggested to be responsible for the elimination of the CA.

Keywords
Contrast agent, polymer, subcellular localization, transmission electron microscope, ultrasound imaging
National Category
Medical Engineering
Research subject
Medical Technology
Identifiers
urn:nbn:se:kth:diva-160905 (URN)10.3233/BSI-140099 (DOI)
Funder
EU, FP7, Seventh Framework Programme, 245572
Note

QC 20150401

Available from: 2015-03-03 Created: 2015-03-03 Last updated: 2017-08-15Bibliographically approved
Nogueira, E., Mangialavori, I. C., Loureiro, A., Azoia, N. G., Sarria, M. P., Nogueira, P., . . . Cavaco-Paulo, A. (2015). Peptide Anchor for Folate-Targeted Liposomal Delivery. Biomacromolecules, 16(9), 2904-2910
Open this publication in new window or tab >>Peptide Anchor for Folate-Targeted Liposomal Delivery
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2015 (English)In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 16, no 9, p. 2904-2910Article in journal (Refereed) Published
Abstract [en]

Specific folate receptors are abundantly overexpressed in chronically activated macrophages and in most cancer cells. Directed folate receptor targeting using liposomes is usually achieved using folate linked to a phospholipid or cholesterol anchor. This link is formed using a large spacer like polyethylene glycol. Here, we report an innovative strategy for targeted liposome delivery that uses a hydrophobic fragment of surfactant protein D linked to folate. Our proposed spacer is a small 4 amino acid residue linker. The peptide conjugate inserts deeply into the lipid bilayer without affecting liposomal integrity, with high stability and specificity. To compare the drug delivery potential of both liposomal targeting systems, we encapsulated the nuclear dye Hoechst 34580. The eventual increase in blue fluorescence would only be detectable upon liposome disruption, leading to specific binding of this dye to DNA. Our delivery system was proven to be more efficient (2-fold) in Caco-2 cells than classic systems where the folate moiety is linked to liposomes by polyethylene glycol.

Place, publisher, year, edition, pages
American Chemical Society (ACS), 2015
Keywords
Surfactant Protein-D, Cyclooxygenase-2 Inhibitor, Inflammatory Response, Carbon-Monoxide, In-Vitro, Celecoxib, Cancer, Cells, Methotrexate, Mechanisms
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Biochemistry and Molecular Biology Cell Biology
Identifiers
urn:nbn:se:kth:diva-174926 (URN)10.1021/acs.biomac.5b00823 (DOI)000361341700037 ()26241560 (PubMedID)2-s2.0-84941584525 (Scopus ID)
Note

QC 20151027

Available from: 2015-10-27 Created: 2015-10-09 Last updated: 2017-12-01Bibliographically approved
Loureiro, A., Nogueira, E., Azoia, N., Sárria, M., Abreu, A., Shimanovich, U., . . . Cavaco-Paulo, A. (2015). Size controlled protein nanoemulsions for active targeting of folate receptor positive cells. Colloids and Surfaces B: Biointerfaces, 135, 90-98
Open this publication in new window or tab >>Size controlled protein nanoemulsions for active targeting of folate receptor positive cells
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2015 (English)In: Colloids and Surfaces B: Biointerfaces, ISSN 0927-7765, E-ISSN 1873-4367, Vol. 135, p. 90-98Article in journal (Refereed) Published
Abstract [en]

Bovine serum albumin (BSA) nanoemulsions were produced by high pressure homogenization with a tri-block copolymer (Poloxamer 407), which presents a central hydrophobic chain of polyoxypropylene (PPO) and two identical lateral hydrophilic chains of polyethylene glycol (PEG). We observed a linear correlation between tri-block copolymer concentration and size - the use of 5. mg/mL of Poloxamer 407 yields nanoemulsions smaller than 100. nm. Molecular dynamics and fluorescent tagging of the tri-block copolymer highlight their mechanistic role on the size of emulsions. This novel method enables the fabrication of highly stable albumin emulsions in the nano-size range, highly desirable for controlled drug delivery. Folic Acid (FA)-tagged protein nanoemulsions were shown to promote specific folate receptor (FR)-mediated targeting in FR positive cells. The novel strategy presented here enables the construction of size controlled, functionalized protein-based nanoemulsions with excellent characteristics for active targeting in cancer therapy.

Place, publisher, year, edition, pages
Elsevier, 2015
National Category
Cell Biology
Identifiers
urn:nbn:se:kth:diva-174582 (URN)10.1016/j.colsurfb.2015.06.073 (DOI)000367410200011 ()2-s2.0-84938412704 (Scopus ID)
Note

QC 20151207

Available from: 2015-12-07 Created: 2015-10-07 Last updated: 2017-12-01Bibliographically approved
Grishenkov, D., Adrian, G., Weitzberg, E., Lundberg, J., Harmark, J., Cerroni, B., . . . Janerot Sjöberg, B. (2015). Ultrasound contrast agent loaded with nitric oxide as a theranostic microdevice: Theranostic contrast agent loaded with nitric oxide. Drug Design, Development and Therapy, 9, 2409-2419
Open this publication in new window or tab >>Ultrasound contrast agent loaded with nitric oxide as a theranostic microdevice: Theranostic contrast agent loaded with nitric oxide
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2015 (English)In: Drug Design, Development and Therapy, ISSN 1177-8881, E-ISSN 1177-8881, Vol. 9, p. 2409-2419Article in journal (Refereed) Published
Abstract [en]

The current study describes novel multifunctional polymer-shelled microbubbles (MBs) loaded with nitric oxide (NO) for integrated therapeutic and diagnostic applications, i.e. theranostics, of myocardial ischemia. We used gas filled MBs with an average diameter of 4 µm stabilized by a biocompatible shell of poly(vinyl)alcohol. In vitro acoustic tests showed a sufficient enhancement of the backscattered power (20 dB) acquired from the MBs suspension. The values of attenuation coefficient (0.8 dB/cm MHz) and phase velocities (1517 m/s) were comparable to those reported for the soft tissue. Moreover, polymer MBs demonstrate increased stability compared to clinically approved contrast agents with fracture threshold of about 900 kPa. In vitro chemiluminescence measurements demonstrated that dry powder of NO-loaded MBs releases its gas content in about 2 hours following an exponential decay profile with an exponential time constant equal 36 min. The application of high power ultrasound pulse (MI=1.2) on the MBs resuspended in saline decreases the exponential time constant from 55 to 4 min in air saturated solution and from 17 to 10 min in degased solution. Thus, ultrasound-triggered release of NO is achieved. Cytotoxicity tests indicate that phagocytosis of the MBs by macrophages starts within 6 to 8 hours. This is suitable time for initial diagnostics, treatment and monitoring of the therapeutic effect using single injection of the proposed multifunctional MBs.

National Category
Medical Equipment Engineering Medical Materials
Identifiers
urn:nbn:se:kth:diva-159765 (URN)10.2147/DDDT.S77790 (DOI)000353631900001 ()2-s2.0-84929178166 (Scopus ID)
Note

QC 20150807. Updated from accepted to published.

Available from: 2015-02-10 Created: 2015-02-10 Last updated: 2017-12-04Bibliographically approved
Poehlmann, M., Grishenkov, D., Kothapalli, S. V. .., Härmark, J., Hebert, H., Philipp, A., . . . Frey, A. (2014). On the interplay of shell structure with low- and high-frequency mechanics of multifunctional magnetic microbubbles. Soft Matter, 10(1), 214-226
Open this publication in new window or tab >>On the interplay of shell structure with low- and high-frequency mechanics of multifunctional magnetic microbubbles
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2014 (English)In: Soft Matter, ISSN 1744-683X, E-ISSN 1744-6848, Vol. 10, no 1, p. 214-226Article in journal (Refereed) Published
Abstract [en]

Polymer-shelled magnetic microbubbles have great potential as hybrid contrast agents for ultrasound and magnetic resonance imaging. In this work, we studied US/MRI contrast agents based on air-filled poly(vinyl alcohol)-shelled microbubbles combined with superparamagnetic iron oxide nanoparticles (SPIONs). The SPIONs are integrated either physically or chemically into the polymeric shell of the microbubbles (MBs). As a result, two different designs of a hybrid contrast agent are obtained. With the physical approach, SPIONs are embedded inside the polymeric shell and with the chemical approach SPIONs are covalently linked to the shell surface. The structural design of hybrid probes is important, because it strongly determines the contrast agent's response in the considered imaging methods. In particular, we were interested how structural differences affect the shell's mechanical properties, which play a key role for the MBs' US imaging performance. Therefore, we thoroughly characterized the MBs' geometric features and investigated low-frequency mechanics by using atomic force microscopy (AFM) and high-frequency mechanics by using acoustic tests. Thus, we were able to quantify the impact of the used SPIONs integration method on the shell's elastic modulus, shear modulus and shear viscosity. In summary, the suggested approach contributes to an improved understanding of structure-property relations in US-active hybrid contrast agents and thus provides the basis for their sustainable development and optimization.

Keywords
Geometric feature, High frequency HF, Imaging performance, Integration method, Physical approaches, Structural differences, Structure property relation, Superparamagnetic iron oxide nanoparticles
National Category
Medical Engineering
Identifiers
urn:nbn:se:kth:diva-134611 (URN)10.1039/c3sm51560e (DOI)000327849000024 ()2-s2.0-84889577207 (Scopus ID)
Funder
EU, FP7, Seventh Framework Programme, 245572
Note

QC 20150626

Available from: 2013-11-25 Created: 2013-11-25 Last updated: 2017-12-06Bibliographically approved
Nogueira, E., Loureiro, A., Nogueira, P., Freitas, J., Almeida, C. R., Härmark, J., . . . Cavaco-Paulo, A. (2013). Liposome and protein based stealth nanoparticles. Faraday discussions (Online), 166, 417-429
Open this publication in new window or tab >>Liposome and protein based stealth nanoparticles
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2013 (English)In: Faraday discussions (Online), ISSN 1359-6640, E-ISSN 1364-5498, Vol. 166, p. 417-429Article in journal (Refereed) Published
Abstract [en]

Liposomes and protein based nanoparticles were tuned with different polymers and glycolipids to improve stealth and thus decrease their clearance by macrophages. Liposomes were coated with polyethylene glycol (PEG) and brain-tissue-derived monosialoganglioside (GM1). Bovine serum albumin (BSA) nanoparticles were produced incorporating a PEGylated surfactant (PEG-surfactant). All obtained nanoparticles were monodisperse, with sizes ranging from 80 to 120 nm, with a zeta-potential close to zero. The presented stealth strategies lead to a decrease of internalization levels by macrophages. These surface modified nanoparticles could be used for production of new drug delivery nanosystems for systemic administration (e.g. intravenous application).

Keywords
Drug-Delivery Systems, Poly(Ethylene Glycol), Circulation Time, In-Vivo, Cancer, Opsonization, Pharmacokinetics, Microspheres, Optimization, Formulations
National Category
Other Chemistry Topics
Identifiers
urn:nbn:se:kth:diva-141099 (URN)10.1039/c3fd00057e (DOI)000329305100026 ()24611291 (PubMedID)2-s2.0-84887481690 (Scopus ID)
Funder
EU, FP7, Seventh Framework Programme, NMP4-LA-2009-228827 NANOFOL
Note

QC 20150623

Available from: 2014-02-07 Created: 2014-02-07 Last updated: 2017-12-06Bibliographically approved
Poehlman, M., Kothapalli, V. V., Grishenkov, D., Härmark, J., Hebert, H., Philipp, A., . . . Fery, A. (2013). Magnetic microbubbles for multimodality imaging: the importance of the shell structure for low and high frequency mechanics. In: : . Paper presented at European symposium and exhibition on biomaterials and related areas, Euro BioMAT2013 23-24 Apr 2013 Weimar, Germany.
Open this publication in new window or tab >>Magnetic microbubbles for multimodality imaging: the importance of the shell structure for low and high frequency mechanics
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2013 (English)Conference paper, Oral presentation with published abstract (Refereed)
Abstract [en]

There is a growing interest in magnetic microbubbles (MBs) for simultaneous enhanced ultrasound (US) and enhanced magnetic resonance imaging (MRI) to support well-established imaging procedures as well as new emerging diagnostic and therapeutic applications. However, the development of hybrid contrast agents is challenging, because their design needs to satisfy a variety of requirements such as a sufficient stability of the probe for the circulation within the cardiovascular system, the production of an adequate US echo signal and a reasonable reduced relaxation time of nearby located protons. The studied magnetic MBs consist of an air-filled core, which is encapsulated by a soft hydrogel-like shell composed of poly(vinyl alcohol) and superparamagnetic iron oxide nanoparticles (SPIONs)[1]. Two strategies were used to combine magnetic nanoparticles with the polymeric shell: SPIONs were either covalently attached to the shell surface via a post-chemical treatment or embedded physically inside the shell during the MBs’ synthesis. In particular, we were interested on the impact of the used SPIONs integration strategy on low and high frequency mechanics of the magnetic MBs. Therefore, we used a straightforward characterization of the MBs on the single particle level to correlate the synthesis with the MBs’ morphological properties and low frequency mechanics that were studied in quasi-static force measurements with atomic force microscopy. High frequency mechanics were investigated by exposure of an ensemble of MBs to an acoustic field. By further correlation of low and high frequency mechanics, we were able to bridge the gap between synthesis and the MBs macroscopic properties relevant for their application. The shown approach offers the possibility to sustainable design and optimize complex probes based on an improved understanding of structure/property relations.

National Category
Medical Laboratory and Measurements Technologies
Identifiers
urn:nbn:se:kth:diva-137644 (URN)
Conference
European symposium and exhibition on biomaterials and related areas, Euro BioMAT2013 23-24 Apr 2013 Weimar, Germany
Note

QC 20140205

Available from: 2013-12-14 Created: 2013-12-14 Last updated: 2016-11-25Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0002-9604-0511

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