RESUMO
Fluorescence imaging is one of the most widely used in vivo imaging methods for both fundamental research and clinical practice. Due to the reduced photon scattering, absorption, and autofluorescence in tissues, the emerging near-infrared (NIR) imaging (650-1700 nm) can afford deep tissue imaging with high spatiotemporal resolution and in vivo report the anatomical structures as well as the physiological activities in a whole-body level. Here, we give a brief introduction to fluorescence imaging in the first NIR (NIR-I, 650-950 nm) and second NIR (NIR-II, 1000-1700 nm) windows, summarize the recently developed NIR fluorophores and their applications in whole-body vascular system imaging, precision cancer theranostics, and regenerative medicine. Finally, the clinical applications and future prospects of in vivo NIR fluorescence imaging are also discussed.
Assuntos
Neoplasias , Imagem Óptica , Corantes Fluorescentes , Humanos , Neoplasias/diagnóstico por imagem , FótonsRESUMO
Pseudomonas putida is potentially used in the bioremediation of heavy metals (HMs). Its response to different HMs in different environments is still not fully understood. This study investigated resistance against 12 kinds of metals by P. putida CZ1 planktonic cells and its biofilm in LB and mineral medium (MM). P. putida CZ1 biofilms have high resistance and accumulation capacity for Cu2+, Zn2+, Pb2+, Fe3+, Mn2+, Al3+ and Ni2+, but less resistance to Co2+, Cd2+, Cr2O72-, Ag+ and Hg2+. Bioï¬lms were 2-8 times more resistant to Cu2+ and Zn2+ than planktonic cells. There was a strong correlation between the P content and the accumulation of Cu2+, Zn2+, Fe3+, Mn2+, Pb2+, Ni2+and Al3+ respectively. Confocal laser scanning microscopy (CLSM) combined with live/dead staining study found that cells in the biofilms can keep viable after 36 h under MIC of Cu2+ or Zn2+ both in LB and MM. When the metal concentration increased, cells can be killed gradually. For Cu2+, Zn2+, Fe3+, Mn2+, Pb2+ and Ni2+, higher resistance was found in MM (2-4 times higher) than in LB and higher accumulation of these metals were also found in MM. P. putida CZ1 biofilm cultured in MM with citric acid as carbon source had stronger resistance and accumulation ability to Cu2+, Zn2+, Pb2+, Fe3+, Mn2+, and Ni2+. This suggested that P. putida CZ1 had greater remediation potential for these metals in organic acid rich environments.
Assuntos
Metais Pesados , Pseudomonas putida , Biodegradação Ambiental , Biofilmes , Monitoramento Ambiental , Metais Pesados/análise , Minerais , Pseudomonas putida/genéticaRESUMO
Accurate intraoperative tissue identification is critical to tumor surgery. However, conventional methods are labor- and time-intensive, which greatly delay the intraoperative decision-making. Herein, a matrix metalloproteinase (MMP)14-activated NIR-II nanoprobe (A&MMP@Ag2 S-AF7P) is presented for rapid unperturbed-tissue analysis for exâ vivo and inâ vivo neuroblastoma diagnosis. A&MMP@Ag2 S-AF7P displays negligible fluorescence in normal tissues but is activated quickly by inhibiting the fluorescence resonance energy transfer (FRET) between Ag2 S QDs and A1094 mediated by MMP14 overexpressed in neuroblastoma; meanwhile, the exposure of the membrane penetrating peptide R9 (TAT-peptide) results in efficient internalization of nanoprobes in the cancer cells, providing superior tumor-to-normal (T/N) tissue ratio. Instant illumination of the lesion and well-defined tumor margins make the nanoprobes a suitable rapid diagnostic reagent for cancer surgical or tissue biopsy procedures.
Assuntos
Raios Infravermelhos/uso terapêutico , Nanopartículas/química , Neoplasias/diagnóstico , HumanosRESUMO
Fluorescence imaging has become a fundamental tool for biomedical applications; nevertheless, its intravital imaging capacity in the conventional wavelength range (400-950 nm) has been restricted by its extremely limited tissue penetration. To tackle this challenge, a novel imaging approach using the fluorescence in the second near-infrared window (NIR-II, 1000-1700 nm) has been developed in the past decade to achieve deep penetration and high-fidelity imaging, and thus significant biomedical applications have begun to emerge. In this Perspective, we first examine recent discoveries and challenges in the development of novel NIR-II fluorophores and compatible imaging apparatuses. Subsequently, the recent advances in bioimaging, biosensing, and therapy using such a cutting-edge imaging technique are highlighted. Finally, based on the achievement in the representative studies, we elucidate the main concerns regarding this imaging technique and give some advice and prospects for the development of NIR-II imaging for future biomedical applications.
Assuntos
Pesquisa Biomédica , Corantes Fluorescentes/química , Imagem Óptica , Animais , Humanos , Raios InfravermelhosRESUMO
Activatable theranostic systems show potential for improved tumor diagnosis and therapy owing to high detection specificities, effective ablation, and minimal side-effects. Herein, a tumor microenvironment (TME)-activated NIR-II nanotheranostic system (FEAD1) for precise diagnosis and treatment of peritoneal metastases is presented. FEAD1 was fabricated by self-assembling the peptide Fmoc-His, mercaptopropionic-functionalized Ag2 S quantum dots (MPA-Ag2 S QDs), the chemodrug doxorubicin (DOX), and NIR absorber A1094 into nanoparticles. We show that in healthy tissue, FEAD1 exists in an NIR-II fluorescence "off" state, because of Ag2 S QDs-A1094 interactions, while DOX remains in stealth mode. Upon delivery of FEAD1 to the tumor, the acidic TME triggers its disassembly through breakage of the Fmoc-His metal coordination and DOX hydrophobic interactions. Release of A1094 switches on Ag2 S fluorescence, illuminating the tumor, accompanied by burst release of DOX within the tumor tissue, thereby achieving precise tumor theranostics. This TME-activated theranostic strategy holds great promise for future clinical applications.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Doxorrubicina/farmacologia , Nanopartículas/química , Neoplasias Peritoneais/diagnóstico por imagem , Neoplasias Peritoneais/tratamento farmacológico , Nanomedicina Teranóstica , Animais , Antibióticos Antineoplásicos/administração & dosagem , Antibióticos Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/administração & dosagem , Doxorrubicina/química , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Concentração de Íons de Hidrogênio , Raios Infravermelhos , Injeções Intraperitoneais , Camundongos , Camundongos Nus , Nanopartículas/administração & dosagem , Neoplasias Experimentais/diagnóstico por imagem , Neoplasias Experimentais/tratamento farmacológico , Imagem Óptica , Pontos Quânticos/administração & dosagem , Pontos Quânticos/química , Compostos de Prata/administração & dosagem , Compostos de Prata/química , Células Tumorais Cultivadas , Microambiente Tumoral/efeitos dos fármacosRESUMO
Traumatic brain injury (TBI) is one of the most dangerous acute diseases resulting in high morbidity and mortality. Current methods remain limited with respect to early diagnosis and real-time feedback on the pathological process. Herein, a targeted activatable fluorescent nanoprobe (V&A@Ag2 S) in the second near-infrared window (NIR-II) is presented for inâ vivo optical imaging of TBI. Initially, the fluorescence of V&A@Ag2 S is turned off owing to energy transfer from Ag2 S to the A1094 chromophore. Upon intravenous injection, V&A@Ag2 S quickly accumulates in the inflamed vascular endothelium of TBI based on VCAM1-mediated endocytosis, after which the nanoprobe achieves rapid recovery of the NIR-II fluorescence of Ag2 S quantum dots (QDs) owing to the bleaching of A1094 by the prodromal biomarker of TBI, peroxynitrite (ONOO- ). The nanoprobe offers high specificity, rapid response, and high sensitivity toward ONOO- , providing a convenient approach for inâ vivo early real-time assessment of TBI.
Assuntos
Lesões Encefálicas Traumáticas/diagnóstico por imagem , Lesões Encefálicas Traumáticas/diagnóstico , Imagem Óptica/métodos , Humanos , NanopartículasRESUMO
Fluorescence-guided cytoreductive surgery is one of the most promising approaches for facile elimination of tumors in situ, thereby improving prognosis. Reported herein is a simple strategy to construct a novel chainlike NIR-II nanoprobe (APP-Ag2 S-RGD) by self-assembly of an amphiphilic peptide (APP) into a nanochain with subsequent chemical crosslinking of NIR-II Ag2 S QDs and the tumor-targeting RGD peptide. This probe exhibits higher capability for cancer cell detection compared with that of RGD-functionalized Ag2 S QDs (Ag2 S-RGD) at the same concentration. Upon intraperitoneal injection, superior tumor-to-normal tissue signal ratio is achieved and non-vascularized tiny tumor metastatic foci as small as about 0.2â mm in diameter could be facilely eliminated under NIR-II fluorescent imaging guidance. These results clearly indicate the potential of this probe for fluorescence-guided tumor staging, preoperative diagnosis, and intraoperative navigation.
Assuntos
Antineoplásicos/química , Materiais Biocompatíveis/química , Corantes Fluorescentes/química , Nanoestruturas/química , Imagem Óptica , Peptídeos/química , Neoplasias Peritoneais/diagnóstico por imagem , Animais , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Corantes Fluorescentes/síntese química , Corantes Fluorescentes/farmacologia , Humanos , Raios Infravermelhos , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C , Neoplasias Experimentais/diagnóstico por imagem , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/cirurgia , Peptídeos/síntese química , Peptídeos/farmacologia , Neoplasias Peritoneais/tratamento farmacológico , Neoplasias Peritoneais/cirurgiaRESUMO
Stem-cell-based regenerative medicine holds great promise in clinical practices. However, the fate of stem cells after transplantation, including the distribution, viability, and the cell clearance, is not fully understood, which is critical to understand the process and the underlying mechanism of regeneration for better therapeutic effects. Herein, we develop a dual-labeling strategy to in situ visualize the fate of transplanted stem cells in vivo by combining the exogenous near-infrared fluorescence imaging in the second window (NIR-II) and endogenous red bioluminescence imaging (BLI). The NIR-II fluorescence of Ag2 S quantum dots is employed to dynamically monitor the trafficking and distribution of all transplanted stem cells in vivo due to its deep tissue penetration and high spatiotemporal resolution, while BLI of red-emitting firefly luciferase (RfLuc) identifies the living stem cells after transplantation in vivo because only the living stem cells express RfLuc. This facile strategy allows for in situ visualization of the dynamic trafficking of stem cells in vivo and the quantitative evaluation of cell translocation and viability with high temporal and spatial resolution, and thus reports the fate of transplanted stem cells and how the living stem cells help, regeneration, for an instance, of a mouse with acute liver failure.
Assuntos
Imagem Óptica/métodos , Células-Tronco/citologia , Animais , Humanos , Falência Hepática/diagnóstico por imagem , Regeneração Hepática/fisiologia , Medições Luminescentes , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pontos QuânticosRESUMO
The low structural stability of hydrogen-bonded organic frameworks (HOFs) is a thorny issue retarding the development of HOFs. A rational design approach is now proposed for construction of a stable HOF. The resultant HOF (PFC-1) exhibits high surface area of 2122â m2 g-1 and excellent chemical stability (intact in concentrated HCl for at least 117â days). A new method of acid-assisted crystalline redemption is used to readily cure the thermal damage to PFC-1. With periodic integration of photoactive pyrene in the robust framework, PFC-1 can efficiently encapsulate Doxorubicin (Doxo) for synergistic chemo-photodynamic therapy, showing comparable therapeutic efficacy with the commercial Doxo yet considerably lower cytotoxicity. This work demonstrates the notorious stability issue of HOFs can be properly addressed through rational design, paving a way to develop robust HOFs and offering promising application perspectives.
Assuntos
Compostos Orgânicos/química , Fotoquimioterapia , Adsorção , Antibióticos Antineoplásicos/química , Cristalização , Doxorrubicina/química , Ligação de Hidrogênio , Microscopia Eletrônica de Varredura , Estrutura Molecular , Difração de Pó , Estudo de Prova de Conceito , TermodinâmicaRESUMO
In this work, a novel type of nanomedical platform, the double-walled Au nanocage/SiO(2) nanorattle, is successfully fabricated by combining two "hollow-excavated strategies"--galvanic replacement and "surface-protected etching". The rational design of double-walled nanostructure based on gold nanocages (AuNCs) and hollow SiO(2) shells functionalized respectively with p-aminothiophenol (pATP) and Tat peptide simultaneously renders the nanoplatforms three functionalities: 1) the whole nanorattle serves as a high efficient drug carrier thanks to the structural characteristics of AuNC and SiO(2) shell with hollow interiors and porous walls; 2) the AuNC with large electromagnetic enhancement acts as a sensitive surface-enhanced Raman scattering (SERS) substrate to track the internalization process of the nanorattles by human MCF-7 breast cancer cells, as well as an efficient photothermal transducer for localized hyperthermia cancer therapy due to the strong near-infrared absorption; 3) Tat-functionalized SiO(2) shell not only improves biocompatibility and cell uptake efficiency resulting in enhanced anticancer efficacy but also prevents the AuNCs from aggregation and provides the stability of AuNCs so that the SERS signals can be used for cell tracking in high fidelity. The reported chemistry and the designed nanostructures should inspire more interesting nanostructures and applications.
Assuntos
Sistemas de Liberação de Medicamentos , Ouro/química , Nanopartículas Metálicas/química , Nanotecnologia/métodos , Fototerapia/métodos , Dióxido de Silício/química , Compostos de Anilina/química , Antineoplásicos/química , Materiais Biocompatíveis/química , Calibragem , Radiação Eletromagnética , Humanos , Concentração de Íons de Hidrogênio , Células MCF-7 , Microscopia Eletrônica de Transmissão , Neoplasias/metabolismo , Fotoquímica , Análise Espectral Raman , Compostos de Sulfidrila/química , Produtos do Gene tat do Vírus da Imunodeficiência Humana/químicaRESUMO
The extracellular polymeric substance (EPS) and surface properties of unsaturated biofilms of a heavy metal-resistant rhizobacterium Pseudomonas putida CZ1, in response to aging, pH, temperature and osmotic stress, were studied by quantitative analysis of EPS and atomic force microscope. It was found that EPS production increased approximately linearly with culture time, cells in the air-biofilm interface enhanced EPS production and decreased cell volume to cope with nutrient depletion during aging. Low pH, high temperature and certain osmotic stress (120 mM NaCl) distinctly stimulated EPS production, and the main component enhanced was extracellular protein. In addition to the enhancement of EPS production in response to high osmotic (328 mM NaCl) stress, cells in the biofilm adhere tightly together to maintain a particular microenvironment. These results indicated the variation of EPS composition and the cooperation of cells in the biofilms is important for the survival of Pseudomonas putida CZ1 from environmental stresses in the unsaturated environments such as rhizosphere.
Assuntos
Biofilmes/crescimento & desenvolvimento , Biopolímeros/metabolismo , Pseudomonas putida/fisiologia , Estresse Fisiológico , Propriedades de Superfície , Aderência Bacteriana , Concentração de Íons de Hidrogênio , Microscopia de Força Atômica , Pressão Osmótica , Pseudomonas putida/efeitos dos fármacos , Pseudomonas putida/efeitos da radiação , Cloreto de Sódio/metabolismo , TemperaturaRESUMO
Intravital fluorescence imaging in the second near-infrared window (NIR-II, 900-1700 nm) has emerged as a promising method for non-invasive diagnostics in complex biological systems due to its advantages of less background interference, high tissue penetration depth, high imaging contrast, and sensitivity. However, traditional NIR-II fluorescence imaging, which is characterized by the "always on" or "turn on" mode, lacks the ability of quantitative detection, leading to low reproducibility and reliability during bio-detection. In contrast, NIR-II ratiometric fluorescence imaging can realize quantitative and reliable analysis and detection in vivo by providing reference signals for fluorescence correction, generating new opportunities and prospects during in vivo bioimaging and biosensing. In this review, the current design strategies and sensing mechanisms of NIR-II ratiometric fluorescence probes for bioimaging and biosensing applications are systematically summarized. Further, current challenges, future perspectives and opportunities for designing NIR-II ratiometric fluorescence probes are also discussed. It is hoped that this review can provide effective guidance for the design of NIR-II ratiometric fluorescence probes and promote its adoption in reliable biological imaging and sensing in vivo.
RESUMO
Stem cell-based cancer treatment has garnered significant attention, yet its safety and efficacy remain incompletely understood. The nuclear factor-kappa B (NF-κB) pathway, a critical signaling mechanism involved in tumor growth, angiogenesis, and invasion, serves as an essential metric for evaluating the behavior of stem cells in tumor models. Herein, we report the development of a triple-channel imaging system capable of simultaneously monitoring the tropism of stem cells towards tumors, assessing tumor proliferation, and quantifying tumor NF-κB activity. In this system, we generated a CRISPR-Cas9 gene-edited human glioblastoma cell line, GE-U87-MG, which provided a reliable readout of the proliferation and NF-κB activity of tumors by EF1α-RFLuc- and NF-κB-GLuc-based bioluminescent imaging, respectively. Additionally, near infrared-II emitting Tat-PEG-AgAuSe quantum dots were developed for tracking of stem cell tropism towards tumor. In a representative case involving human mesenchymal stem cells (hMSCs), multichannel imaging revealed no discernible effect of hMSCs on the proliferation and NF-κB activity of GE-U87-MG tumors. Moreover, hMSCs engineered to overexpress the necrosis factor-related apoptosis-inducing ligand were able to inhibit NF-κB activity and growth of GE-U87-MG in vivo. Taken together, our imaging system represents a powerful and feasible approach to evaluating the safety and therapeutic efficacy of stem cells in tumor models.
Assuntos
Glioblastoma , NF-kappa B , Humanos , NF-kappa B/metabolismo , Linhagem Celular Tumoral , Células-Tronco/metabolismo , Apoptose , Glioblastoma/diagnóstico por imagem , Glioblastoma/terapia , Glioblastoma/metabolismo , Proliferação de Células , Imagem Óptica , TropismoRESUMO
In order to investigate the microbe-mineral interaction in the micro scale, spatial distribution and speciation of Cu and S in Halothiobacillus HT1 biofilm formed on a CuS surface was examined using synchrotron-based X-ray techniques. Confocal laser scanning microscope (CLSM) results indicated that Halothiobacillus HT1 biofilm formation gave rise to distinct chemical and redox gradients, leading to diverse niches in the biofilm. Live cells were distributed at the air-biofilm and membrane-biofilm interface. CuS was oxidized by Halothiobacillus HT1 biofilm, and copper penetrated into the biofilm. Sulfide was oxidized to cysteine (77.3%), sulfite (3.8%) and sulfonate (18.9%). Cu-cysteine-like species were involved in the copper homeostasis. These results significantly improve our understanding of the interfacial properties of the biofilm-mineral interface.
Assuntos
Biofilmes/efeitos dos fármacos , Cobre/farmacologia , Halothiobacillus/citologia , Halothiobacillus/fisiologia , Síncrotrons , Halothiobacillus/efeitos dos fármacos , Viabilidade Microbiana/efeitos dos fármacos , Microscopia de Fluorescência , Enxofre/isolamento & purificação , Espectroscopia por Absorção de Raios X , Raios XRESUMO
Early monitoring of gastrointestinal diseases via orally delivered NIR-II ratiometric fluorescent probes represents a promising noninvasive diagnostic modality, but is challenging due to the limitation of harsh digestive environment. Here, we report a single-component NIR-II ratiometric molecular nanoprobe (LC-1250 NP) to monitor gastrointestinal disease with high specificity to its biomarker H2O2 via oral administration. LC-1250 NP displays stable fluorescence in the channel of 1250 long-pass (F1250LP) before and after the gastrointestinal disease detection as the reference, while it presents significantly enhanced fluorescence signal in the response channel of 1150 nm short-pass (F1150SP) in diseased gastrointestinal environment due to the intramolecular cyclization of LC-1250 molecules activated by H2O2. The fluorescence ratio (F1150SP/F1250LP) increases linearly with the concentration of H2O2 with a low detection limit of 20 nM. Therefore, when delivered orally, LC-1250 NP can accurately map the diseased areas and surmount the false-positive interference from biological heterogeneity by NIR-II ratiometric fluorescence imaging, providing sensitive and reliable evaluation for the progress of gastroenteritis.
Assuntos
Corantes Fluorescentes , Gastroenteropatias , Humanos , Peróxido de Hidrogênio , Nanotecnologia , FluorescênciaRESUMO
Effective drug delivery in the central nervous system (CNS) needs to have long blood-circulation half-lives, to pass through the blood-brain barrier (BBB), and subsequently to be taken up by target cells. Herein, a traceable CNS delivery nanoformulation (RVG-NV-NPs) is developed by encapsulating bexarotene (Bex) and AgAuSe quantum dots (QDs) within Lamp2b-RVG-overexpressed neural stem cell (NSC) membranes. The high-fidelity near-infrared-II imaging by AgAuSe QDs offers a possibility of in vivo monitoring the multiscale delivery process of the nanoformulation from the whole-body to the single-cell scale. It was revealed the synergy of acetylcholine receptor-targeting of RVG and the natural brain-homing and low immunogenicity of NSC membranes prolong the blood circulation, facilitate BBB crossing and nerve cell targeting of RVG-NV-NPs. Thus, in Alzheimer's disease (AD) mice, the intravenous delivery of as low as 0.5% of oral dose Bex showed highly effective up-regulation of the apolipoprotein E expression, resulting rapid alleviation of â¼40% ß-amyloid (Aß) level in the brain interstitial fluid after a single dose administration. The pathological progression of Aß in AD mice is completely suppressed during a 1 month treatment, thus effectively protecting neurons from Aß-induced apoptosis and maintaining the cognitive abilities of AD mice.
Assuntos
Doença de Alzheimer , Camundongos , Animais , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/tratamento farmacológico , Encéfalo/metabolismo , Barreira Hematoencefálica/metabolismo , Peptídeos beta-Amiloides/metabolismo , Membrana Celular/metabolismoRESUMO
Precise drug delivery in cancer treatment is a long-standing concern of modern medicine. Compared with traditional molecular medicines and nano-medicines, emerging cell-based biomimetic delivery strategies display numerous merits, including successive biological functions, innate biocompatibility and superior security since they originate from living organisms, providing a very promising approach. Among them, immune cells receive increasing attention because of their inherent ability in tumor resistance, pathogen elimination, and other significant physiological functions. Herein, we investigated the recent advances on immune cell-based high efficient delivery and therapeutic strategies in solid tumor treatment, mainly focus on T cells, natural killer cells and macrophages, which have been used as drug cargos directly or provided membrane/exosomes as nanoscale drug delivery systems. We also discuss the further potential applications and perspective of this innovative strategy, as well as the predictable challenges in forward exploration in this emerging area.
Assuntos
Bioengenharia , Materiais Biomiméticos , Sistemas de Liberação de Medicamentos , Neoplasias , Animais , Humanos , Imunoterapia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Neoplasias/imunologia , Neoplasias/terapia , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Controllable regulation of stem cell differentiation is a critical concern in stem cell-based regenerative medicine. In particular, there are still great challenges in controlling the directional differentiation of neural stem cells (NSCs) into neurons. Herein, we developed a novel linear-branched poly(ß-amino esters) (S4-TMPTA-BDA-DT, STBD) through a two-step reaction. The synthesized linear-branched polymers possess multiple positively charged amine terminus and degradable intermolecular ester bonds, thus endowing them with excellent properties such as high gene load, efficient gene delivery, and effective gene release and transcription in cells. In the mCherry transfection test, a high transfection efficiency of approximately 70% was achieved in primary NSCs after a single transfection. Moreover, STBD also showed high biocompatibility to NSCs without disturbing their viability and neural differentiation. With the high gene delivery property, STBD is capable of delivering siRNA (shSOX9) expression plasmid into NSCs to significantly interfere with the expression of SOX9, thus enhancing the neuronal differentiation and maturation of NSCs. The STBD/DNA nano-polyplex represents a powerful non-viral approach of gene delivery for manipulating the differentiation of stem cells, showing broad application prospects in NSC-based regenerative therapy for treating neurodegenerative diseases.
Assuntos
Ésteres , Células-Tronco Neurais , Diferenciação Celular/genética , DNA/química , TransfecçãoRESUMO
Axonal transport plays a significant role in the establishment of neuronal polarity, axon growth, and synapse formation during neuronal development. The axon of a naturally growing neuron is a highly complex and multifurcated structure with a large number of bends and branches. Nowadays, the study of dynamic axonal transport in morphologically complex neurons is greatly limited by the technological barrier. Here, a sparse gene transfection strategy was developed to locate fluorescent mCherry in the lysosome of primary neurons, thus enabling us to track the lysosome-based axonal transport with a single-particle resolution. Thereby, several axonal transport models were observed, including the forward or backward transport model, stop-and-go model, repeated back-and-forth transport model, and cross-branch transport model. Then, the accurate single-particle velocity quantification by TrackMate revealed a highly heterogeneous and discontinuous transportation process of lysosome-based axonal transport in freely orientated axons. And, multiple physical factors, such as the axonal structure and the size of particles, were disclosed to affect the velocity of particle transporting in freely orientated axons. The combined single-particle fluorescence tracking and TrackMate assay can be served as a facile tool for evaluating axonal transport in neuronal development and axonal transport-related diseases.
Assuntos
Transporte Axonal , Axônios , Transporte Axonal/fisiologia , Axônios/metabolismo , Fluorescência , Lisossomos , NeurôniosRESUMO
The spatial and temporal distribution of metals in unsaturated Pseudomonas putida CZ1 biofilms was determined using synchrotron-based X-ray fluorescence microscopy (XRF). It was found that Fe, Mn, and Ca were mainly distributed near the air-biofilm interface of a biofilm grown on 40 mM citrate, while there were two Fe-, Mn-, and Ca-rich layers within a biofilm grown on 10 mM citrate. The sorption of copper by biofilm grown in medium containing 10 mM citrate was rapid, with copper being found throughout the biofilm after only 1 h of exposure. Copper initially colocalized with Fe and Mn element layers in the biofilm and then precipitated in a 40-µm-thick layer near the air-biofilm interface when exposed for 12 h. Cu K-edge X-ray absorption near edge structure (XANES) analysis revealed that Cu was primarily bound with citrate within the biofilm, and the precipitate formed in the biofilm exposed to copper for 12 h was most similar to copper phosphate. LIVE/DEAD staining revealed that cells at the biofilm-membrane interface were mostly alive even when the copper concentration reached 80.5 mg copper g(-1) biomass. This suggests that the biofilm matrix provided significant protection for cells in this area. These results significantly improve our understanding of metal acquisition, transportation, and immobilization in unsaturated biofilm systems.