Multifunctional nanoparticles for hyperthermia, thermometry and fluorescenceimaging in the biological windows

Tesis Doctoral inédita cotutelada por la Universidade Federal de Alagoas de Brasil y la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Física de Materiales. Fecha de lectura: 17-12-2018In this thesis, the use of core/shell engineering for the synthesis of fluorescent nanoparticles (NPs) capable of operating as nanothermometers, nanoheaters and/or contrast agents for fluorescence imaging in small animal models is explored. The materials here studied – rare-earth (Nd3+, Yb3+, Tm3+ and/or Er3+) doped NPs and PbS/CdS/ZnS quantum dots (QDs) – presented emission and/or excitation bands in the so-called biological windows, where light penetration into tissues is maximal, allowing for ex vivo and in vivo applications. It was demonstrated that the spatial separation between the rare-earth ions, achieved by the core/shell nano-engineering, resulted not only in a considerable improvement on the values of thermo-optical parameters such as the light-heat conversion efficiency and the relative thermal sensitivity, but also on a multi-functionality of the nanosystems. As a consequence, innovative applications in nanothermometry were successfully accomplished when developing this thesis. Among those applications, one can mention: the study in real time of the thermal dynamics of an in vivo tissue, the detection and monitoring of cardiovascular diseases and the recording of in vivo thermal images and videos at a subcutaneous level by means of a ratiometric approach. The results here presented open up avenues for new diagnosis and control techniques that can revolutionize the current methods found in biomedicin

New opportunities for light-based tumor treatment with an “iron fist”

The efficacy of photodynamic treatments of tumors can be significantly improved by using a new generation of nanoparticles that take advantage of the unique properties of the tumor microenvironmen

The role of tissue fluorescence in in vivo optical bioimaging

The following article appeared in Journal of Applied Physics 128.17 (2020): 171101 and may be found at https://doi.org/10.1063/5.0021854The technological advancements made in optics and semiconductors (e.g., cameras and laser diodes) working with infrared have brought interest in optical bioimaging back to the forefront of research investigating in vivo medical imaging techniques. The definition of the near-infrared transparency windows has turned optical imaging into more than just a method for topical imaging applications. Moreover, this has focused attention back to tissue fluorescence, emissions by tissues and organs that occur when excited by external illumination sources. Most endogenous fluorophores emit in the blue to green range of the electromagnetic spectrum and the resulting tissue fluorescence can be employed in studies from cells to tissue metabolism or avoided by shifting to the red if seen as unwanted autofluorescence. With the more recent move to infrared, it was discovered that autofluorescence is not limited to the visible but also strongly affects in vivo imaging in the infrared. In this Tutorial, we give an overview on tissue fluorescence and tissue interactions with excitation light as well as their effect on in vivo imaging. Furthermore, potential sources of tissue fluorescence in the near-infrared are identified and we describe approaches for successful biomedical imaging in the biological windows, taking into consideration infrared autofluorescence and summarizing techniques for avoiding it in in vivo imaging experimentsThis work was supported by the Spanish Ministry of Economy and Competitiveness under Project No. MAT2016-75362-C3-1-R, the Spanish Ministry of Sciences, Innovation and Universities under Project No. PID2019-106211RB-I00 (NANONERV), by the Instituto de Salud Carlos III (Nos. PI16/00812 and PI19/00565), and through the Comunidad Autónoma de Madrid (No. B2017/ BMD-3867RENIMCM), and co-financed by the European Structural and investment fund. Additional funding was provided by the European Union’s Horizon 2020 FET Open project NanoTBTech (Grant Agreement No. 801305), the Fundación para la Investigación Biomédica del Hospital Universitario Ramón y Cajal under Project No. IMP18_38(2018/0265), and also COST action CA17140. Y.S. acknowledges a scholarship from the China Scholarship Council (No.201806870023), E.X. is grateful for a Juan de la Cierva Formación scholarship (No. FJC2018-036734-I), and D.H.O. is thankful to the Instituto de Salud Carlos III for a Sara Borrell Fellowship (No. CD17/00210). The authors thank Dr. Blanca del Rosal for the helpful discussion and input on the manuscrip

Luminescence based temperature bio-imaging: Status, challenges, and perspectives

The only way to get thermal images of living organisms without perturbing them is to use luminescent probes with temperature-dependent spectral properties. The acquisition of such thermal images becomes essential to distinguish various states of cells, to monitor thermogenesis, to study cellular activity, and to control hyperthermia therapy. Current efforts are focused on the development and optimization of luminescent reporters such as small molecules, proteins, quantum dots, and lanthanide-doped nanoparticles. However, much less attention is devoted to the methods and technologies that are required to image temperature distribution at both in vitro or in vivo levels. Indeed, rare examples can be found in the scientific literature showing technologies and materials capable of providing reliable 2D thermal images of living organisms. In this review article, examples of 2D luminescence thermometry are presented alongside new possibilities and directions that should be followed to achieve the required level of simplicity and reliability that ensure their future implementation at the clinical level. This review will inspire specialists in chemistry, physics, biology, medicine, and engineering to collaborate with materials scientists to jointly develop novel more accurate temperature probes and enable mapping of temperature with simplified technical mean

Luminescence thermometry for brain activity monitoring: A perspective

Minimally invasive monitoring of brain activity is essential not only to gain understanding on the working principles of the brain, but also for the development of new diagnostic tools. In this perspective we describe how brain thermometry could be an alternative to conventional methods (e.g., magnetic resonance or nuclear medicine) for the acquisition of thermal images of the brain with enough spatial and temperature resolution to track brain activity in minimally perturbed animals. We focus on the latest advances in transcranial luminescence thermometry introducing a critical discussion on its advantages and shortcomings. We also anticipate the main challenges that the application of luminescent nanoparticles for brain thermometry will face in next years. With this work we aim to promote the development of near infrared luminescence for brain activity monitoring, which could also benefit other research areas dealing with the brain and its illnessesThis work was financed by the Spanish Ministerio de Innovación y Ciencias under project NANONERV PID 2019-106211RB-I00. BD acknowledges support from the Australian Research Council (DE200100985), RMIT University (Vice-Chancellor’s Fellowship Programme) and the Australian Academy of Sciences (JG Russell Award). PR-S is grateful for a Juan de la Cierva—Incorporación scholarship (IJC2019-041915-I). AB acknowledges funding from Comunidad de Madrid through TALENTO grant ref. 2019-T1/IND-14014. EX is grateful for a Juan de la Cierva - Incorporación scholarship (IJC2020-045229-I

A brighter era for silver chalcogenide semiconductor nanocrystals

Silver chalcogenide semiconductor nanocrystals (Ag2E SNCs) have become a household name in the biomedical field, where they are used as contrast agents in bioimaging, photothermal therapy agents, and luminescent nanothermometers. The prominent position they have come to occupy in this field stems from a unique combination of features, above all near-infrared excitation and emission alongside low cytotoxicity. However, the first reports on Ag2E SNCs showed that a great limitation of these luminescent nanomaterials resided in their low photoluminescence quantum yield, which results in reduced brightness: a crippling feature in bioimaging and biosensing. In this article, we provide an overview of the strategies developed to overcome this hurdle. These strategies aim to remedy the presence of defects in the SNC core and/or surface, the presence of metallic silver, and off-stoichiometric composition. These features stem from the high mobility and redox potential of Ag+ ions, alongside the difficulty in controlling the nucleation and growth rate of Ag2E SNCs. The effectiveness of each approach is discussed. Lastly, a perspective on future research efforts to make Ag2E SNCs even brighter – and thus more effective in biomedical applications – is provided, with the hope of inspiring further investigation on these nanomaterials with a rich, complex set of physicochemical and spectroscopic propertiesThis work was financed by the Spanish Ministerio de Ciencia e Innovacion under project NANONERV PID2019-106211RB-I00, NANOGRANZ PID2021-123318OB-I00, PID2021-122806OB-I00 and TED2021-132317-I00B, by the Instituto de Salud Carlos III (PI19/ 00565), by the Comunidad Autonoma de Madrid (P2022/BMD-7403 RENIM-CM) and co-financed by the European structural and investment fund. R.M. is grateful to the Spanish Ministerio de Ciencia e Innovación for support to research through a Ramón y Cajal Fellowship (RYC2021- 032913-I). I.Z.-G. thanks UCM-Santander for a predoctoral contract (CT63/19-CT64/19). L.M. acknowledges a scholarship from the China Scholarship Council (No. 202108350018

The near-infrared autofluorescence fingerprint of the brain

This is the peer reviewed version of the following article: Lifante, J, del Rosal, B, Chaves-Coira, I, Fernández, N, Jaque, D, Ximendes, E. The near-infrared autofluorescence fingerprint of the brain. J. Biophotonics. 2020; 13:e202000154, which has been published in final form at https://doi.org/10.1002/jbio.202000154. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived VersionsThe brain is a vital organ involved in mostof the central nervous system disorders.Their diagnosis and treatment require fast,cost-effective, high-resolution and high-sensitivity imaging. The combinationof a new generation of luminescent nanoparticles and imaging systems work-ing in the second biological window (near-infrared II [NIR-II]) is emerging asa reliable alternative. For NIR-II imaging to become a robust technique at thepreclinical level, full knowledge of the NIR-II brain autofluorescence, responsi-ble for the loss of image resolution and contrast, is required. This work demon-strates that the brain shows a peculiar infrared autofluorescence spectrumthat can be correlated with specific molecular components. The existence ofparticular structures within the brain with well-defined NIR autofluorescencefingerprints is also evidenced, opening the door to in vivo anatomical imaging.Finally, we propose a rational selection of NIR luminescent probes suitable forlow-noise brain imaging based on their spectral overlap with brainautofluorescenceComunidad de Madrid, Grant/AwardNumber: B2017/BMD-3867RENIMCM;European Cooperation in Science andTechnology, Grant/Award Number:CA17140; Fundación para la Investigación Biomédica del Hospital Universitario Ramón y Cajal, Grant/Award Number:IMP18_38(2018/0265); Horizon 2020 Framework Programme, Grant/AwardNumber: 801305; Instituto de Salud CarlosIII, Grant/Award Number: PI16/00812;Ministerio de Ciencia, Innovación y Universidades, Grant/Award Number:FJC2018-036734-I; Ministerio deEconomía y Competitividad, Grant/AwardNumbers: MAT2016-75362-C3-1-R,MAT2017-83111R, MAT2017-85617-

Neural networks push the limits of luminescence lifetime nanosensing

Luminescence lifetime-based sensing is ideally suited to monitor biological systems due to its minimal invasiveness and remote working principle. Yet, its applicability is limited in conditions of low signal-to-noise ratio (SNR) induced by, e.g., short exposure times and presence of opaque tissues. Herein this limitation is overcome by applying a U-shaped convolutional neural network (U-NET) to improve luminescence lifetime estimation under conditions of extremely low SNR. Specifically, the prowess of the U-NET is showcased in the context of luminescence lifetime thermometry, achieving more precise thermal readouts using Ag2S nanothermometers. Compared to traditional analysis methods of decay curve fitting and integration, the U-NET can extract average lifetimes more precisely and consistently regardless of the SNR value. The improvement achieved in the sensing performance using the U-NET is demonstrated with two experiments characterized by extreme measurement conditions: thermal monitoring of free-falling droplets, and monitoring of thermal transients in suspended droplets through an opaque medium. These results broaden the applicability of luminescence lifetime-based sensing in fields including in vivo experimentation and microfluidics, while, hopefully, spurring further research on the implementation of machine learning (ML) in luminescence sensingThis work was financed by the Spanish Ministerio de Innovación y Ciencias under Project Nos. RTI2018-101050-J-I00, NANONERV PID2019-106211RB-I00, NANOGRANZ PID2021-123318OB-I00, TED2021-132317- I00B, and EIN2020-112419. Additional funding was provided by the European Union Horizon 2020 FETOpen project NanoTBTech (Grant No. 801305) and by the Comunidad Autónoma de Madrid (S2022/BMD7403 REMIN-CM). R.M. is grateful to the Spanish Ministerio de Ciencia e Innovación for support to research through a Ramón y Cajal Fellowship (RYC2021-032913-I). L.M. acknowledges a scholarship from the China Scholarship Council (No. 202108350018). I.Z.-G. thanks UCM-Santander for a predoctoral contract (CT63/19-CT64/19). P.R.-S. is grateful for a Juan de la Cierva-Incorporación scholarship (Grant No. IJC2019-041915-I

In vivo grading of lipids in fatty liver by near-infrared autofluorescence and reflectance

The prevalence of nonalcoholic fatty liver (NAFLD) is rapidly increasing worldwide. When untreated, it may lead to complications such as liver cirrhosis or hepatocarcinoma. The diagnosis of NAFLD is usually obtained by ultrasonography, a technique that can underestimate its prevalence. For this reason, physicians aspire for an accurate, cost-effective, and noninvasive method to determine both the presence and the specific stage of the NAFLD. In this paper, we report an integrated approach for the quantitative estimation of the density of triglycerides in the liver based on the use of autofluorescence and reflectance signals generated by the abdomen of obese C57BL6/J mice. Singular value decomposition is applied to the generated spectra and its corresponding regression model provided a determination coefficient of 0.99 and a root mean square error of 240 mg/dl. This, in turn, enabled the quantitative imaging of triglycerides density in the livers of mice under in vivo conditionsMinisterio de Ciencia e Innovacion, Grant/Award Number: IJC2020-045229-I; Ministerio de Ciencia e Innovacion, Grant/Award Number: NANONERVPID2019-106211RB-I0

Boosting the near-infrared emission of Ag2S nanoparticles by a controllable surface treatment for bioimaging applications

Ag2S nanoparticles are the staple for high-resolution preclinical imaging and sensing owing to their photochemical stability, low toxicity, and photoluminescence (PL) in the second near-infrared biological window. Unfortunately, Ag2S nanoparticles exhibit a low PL efficiency attributed to their defective surface chemistry, which curbs their translation into the clinics. To address this shortcoming, we present a simple methodology that allows to improve the PL quantum yield from 2 to 10%, which is accompanied by a PL lifetime lengthening from 0.7 to 3.8 μs. Elemental mapping and X-ray photoelectron spectroscopy indicate that the PL enhancement is related to the partial removal of sulfur atoms from the nanoparticle's surface, reducing surface traps responsible for nonradiative de-excitation processes. This interpretation is further backed by theoretical modeling. The acquired knowledge about the nanoparticles' surface chemistry is used to optimize the procedure to transfer the nanoparticles into aqueous media, obtaining water-dispersible Ag2S nanoparticles that maintain excellent PL properties. Finally, we compare the performance of these nanoparticles with other near-infrared luminescent probes in a set of in vitro and in vivo experiments, which demonstrates not only their cytocompatibility but also their superb optical properties when they are used in vivo, affording higher resolution image