Complete surgical resection of neoplasia remains one of the most efficient tumor therapies. However, malignant cell clusters are often left behind during surgery due to the inability to visualize and differentiate them against host tissue. Here we establish the feasibility of multicolor fluorescent intravital live microscopy (FILM) where multiple cellular and/or unique tissue compartments are stained simultaneously and imaged in real time.
Methodology/Principal Findings
Theoretical simulations of imaging probe localization were carried out for three agents with specificity for cancer cells, stromal host response, or vascular perfusion. This transport analysis gave insight into the probe pharmacokinetics and tissue distribution, facilitating the experimental design and allowing predictions to be made about the localization of the probes in other animal models and in the clinic. The imaging probes were administered systemically at optimal time points based on the simulations, and the multicolor FILM images obtained in vivo were then compared to conventional pathological sections. Our data show the feasibility of real time in vivo pathology at cellular resolution and molecular specificity with excellent agreement between intravital and traditional in vitro immunohistochemistry.
Conclusions/Significance
Multicolor FILM is an accurate method for identifying malignant tissue and cells in vivo. The imaging probes distributed in a manner similar to predictions based on transport principles, and these models can be used to design future probes and experiments. FILM can provide critical real time feedback and should be a useful tool for more effective and complete cancer resection. 相似文献
The role of lymphatic vessels in tissue and organ transplantation as well as in tumor growth and metastasis has drawn great attention in recent years.
Methodology/Principal Findings
We now developed a novel method using non-invasive two-photon microscopy to simultaneously visualize and track specifically stained lymphatic vessels and autofluorescent adjacent tissues such as collagen fibrils, blood vessels and immune cells in the mouse model of corneal neovascularization in vivo. The mouse cornea serves as an ideal tissue for this technique due to its easy accessibility and its inducible and modifiable state of pathological hem- and lymphvascularization.Neovascularization was induced by suture placement in corneas of Balb/C mice. Two weeks after treatment, lymphatic vessels were stained intravital by intrastromal injection of a fluorescently labeled LYVE-1 antibody and the corneas were evaluated in vivo by two-photon microscopy (TPM). Intravital TPM was performed at 710 nm and 826 nm excitation wavelengths to detect immunofluorescence and tissue autofluorescence using a custom made animal holder. Corneas were then harvested, fixed and analyzed by histology.Time lapse imaging demonstrated the first in vivo evidence of immune cell migration into lymphatic vessels and luminal transport of individual cells. Cells immigrated within 1–5.5 min into the vessel lumen. Mean velocities of intrastromal corneal immune cells were around 9 µm/min and therefore comparable to those of T-cells and macrophages in other mucosal surfaces.
Conclusions
To our knowledge we here demonstrate for the first time the intravital real-time transmigration of immune cells into lymphatic vessels. Overall this study demonstrates the valuable use of intravital autofluorescence two-photon microscopy in the model of suture-induced corneal vascularizations to study interactions of immune and subsequently tumor cells with lymphatic vessels under close as possible physiological conditions. 相似文献
Pancreatic ductal adenocarcinoma (PDAC) carries an extremely poor prognosis, typically presenting with metastasis at the time of diagnosis and exhibiting profound resistance to existing therapies. The development of molecular markers and imaging probes for incipient PDAC would enable earlier detection and guide the development of interventive therapies. Here we sought to identify novel molecular markers and to test their potential as targeted imaging agents.
Methods and Findings
Here, a phage display approach was used in a mouse model of PDAC to screen for peptides that specifically bind to cell surface antigens on PDAC cells. These screens yielded a motif that distinguishes PDAC cells from normal pancreatic duct cells in vitro, which, upon proteomics analysis, identified plectin-1 as a novel biomarker of PDAC. To assess their utility for in vivo imaging, the plectin-1 targeted peptides (PTP) were conjugated to magnetofluorescent nanoparticles. In conjunction with intravital confocal microscopy and MRI, these nanoparticles enabled detection of small PDAC and precursor lesions in engineered mouse models.
Conclusions
Our approach exploited a well-defined model of PDAC, enabling rapid identification and validation of PTP. The developed specific imaging probe, along with the discovery of plectin-1 as a novel biomarker, may have clinical utility in the diagnosis and management of PDAC in humans. 相似文献
While zebrafish embryos are amenable to in vivo imaging, allowing the study of morphogenetic processes during development, intravital imaging of adults is hampered by their small size and loss of transparency. The use of adult zebrafish as a vertebrate model of cardiac disease and regeneration is increasing at high speed. It is therefore of great importance to establish appropriate and robust methods to measure cardiac function parameters.
Methods and Results
Here we describe the use of 2D-echocardiography to study the fractional volume shortening and segmental wall motion of the ventricle. Our data show that 2D-echocardiography can be used to evaluate cardiac injury and also to study recovery of cardiac function. Interestingly, our results show that while global systolic function recovered following cardiac cryoinjury, ventricular wall motion was only partially restored.
Conclusion
Cryoinjury leads to long-lasting impairment of cardiac contraction, partially mimicking the consequences of myocardial infarction in humans. Functional assessment of heart regeneration by echocardiography allows a deeper understanding of the mechanisms of cardiac regeneration and has the advantage of being easily transferable to other cardiovascular zebrafish disease models. 相似文献
Neutrophils are known to be key players in innate immunity. Activated neutrophils induce local inflammation, which results in pathophysiologic changes during intestinal ischemia-reperfusion injury (IRI). However, most studies have been based on static assessments, and few have examined real-time intravital neutrophil recruitment. We herein report a method for imaging and evaluating dynamic changes in the neutrophil recruitment in intestinal IRI using two-photon laser scanning microscopy (TPLSM).
Methods
LysM-eGFP mice were subjected to 45?min of warm intestinal ischemia followed by reperfusion. Mice received an intravenous injection of tetramethylrhodamine isothiocyanate-labeled albumin to visualize the microvasculature. Using a time-lapse TPLSM technique, we directly observed the behavior of neutrophils in intestinal IRI.
Results
We were able to image all layers of the intestine without invasive surgical stress. At low-magnification, the number of neutrophils per field of view continued to increase for 4?h after reperfusion. High-magnification images revealed the presence or absence of blood circulation. At 0–2?h after reperfusion, rolling and adhesive neutrophils increased along the vasculature. At 2–4?h after reperfusion, the irregularity of crypt architecture and transmigration of neutrophils were observed in the lamina propria. Furthermore, TPLSM imaging revealed the villus height, the diameters of the crypt, and the number of infiltrating neutrophils in the crypt. In the IRI group, the villus height 4?h after reperfusion was significantly shorter than in the control group.
Conclusions
TPLSM imaging revealed the real-time neutrophil recruitment in intestinal IRI. Z-stack imaging was useful for evaluating pathophysiological changes in the intestinal wall. 相似文献
The purpose of this paper is to characterize the environmental impacts of equipment used in power transmission and distribution.
This study is divided in two parts, each addressing different main components of the electrical grid system. This part is
concerned with the impacts of transformers and substation equipment while in part 1 a similar analysis is presented for power
lines and cables. 相似文献
Advances in optical imaging modalities and the continued evolution of genetically-encoded fluorescent proteins are coming
together to facilitate the study of cell behavior at high resolution in living organisms. As a result, imaging using autofluorescent
protein reporters is gaining popularity in mouse transgenic and targeted mutagenesis applications. 相似文献
Video cameras sense passively from a distance, offer a rich information stream, and provide intuitively meaningful raw data.
Camera-based imaging has thus proven critical for many advances in neuroscience and biology, with applications ranging from
cellular imaging of fluorescent dyes to tracking of whole-animal behavior at ecologically relevant spatial scales. 相似文献
Pre-operative imaging devices generate high-resolution images but intra-operative imaging devices generate low-resolution
images. To use high-resolution pre-operative images during surgery, they must be deformed to reflect intra-operative geometry
of brain. 相似文献
Optical imaging is an attractive non-invasive way to evaluate the expression of a transferred DNA, mainly thanks to its lower
cost and ease of realization. In this study optical imaging was evaluated for monitoring and quantification of the mouse knee
joint and tibial cranial muscle electrotransfer of a luciferase encoding plasmid. Optical imaging was applied to study the
kinetics of luciferase expression in both tissues. 相似文献
Despite significant progress in imaging technologies, the efficient detection of edges and elongated features in images of
intracellular and multicellular structures acquired using light or electron microscopy is a challenging and time consuming
task in many laboratories. 相似文献
The study was conducted to investigate the diagnostic performance of infrared (IR) imaging of the breast using an interpretive
model derived from a scoring system. 相似文献
Two-photon dual-color imaging of tissues and cells labeled with fluorescent proteins (FPs) is challenging because most two-photon
microscopes only provide one laser excitation wavelength at a time. At present, methods for two-photon dual-color imaging
are limited due to the requirement of large differences in Stokes shifts between the FPs used and their low two-photon absorption
(2PA) efficiency. 相似文献
Different imaging systems (e.g. electrical, magnetic, and ultrasound) rely on a wide variety of physical properties, and the
datasets obtained from such systems provide only partial information about the unknown true state. One approach is to choose
complementary imaging systems, and to combine the information to achieve a better representation. 相似文献
Accurate measurement of peripheral tissue perfusion is challenging but necessary to diagnose peripheral vascular insufficiency. Because near infrared (NIR) radiation can penetrate relatively deep into tissue, significant attention has been given to intravital NIR fluorescence imaging.
Methodology/Principal Findings
We developed a new optical imaging-based strategy for quantitative measurement of peripheral tissue perfusion by time-series analysis of local pharmacokinetics of the NIR fluorophore, indocyanine green (ICG). Time-series NIR fluorescence images were obtained after injecting ICG intravenously in a murine hindlimb ischemia model. Mathematical modeling and computational simulations were used for translating time-series ICG images into quantitative pixel perfusion rates and a perfusion map. We could successfully predict the prognosis of ischemic hindlimbs based on the perfusion profiles obtained immediately after surgery, which were dependent on the preexisting collaterals. This method also reflected increases in perfusion and improvements in prognosis of ischemic hindlimbs induced by treatment with vascular endothelial growth factor and COMP-angiopoietin-1.
Conclusions/Significance
We propose that this novel NIR-imaging-based strategy is a powerful tool for biomedical studies related to the evaluation of therapeutic interventions directed at stimulating angiogenesis. 相似文献
The purpose of this work was to establish and validate combined small animal positron emission tomography - computed tomography
(PET/CT) as a new in vivo imaging method for visualisation and quantification of joint inflammation. 相似文献
This review is aimed at interpreting development and advantages of intravital imaging as an emerging invaluable methodology and summarizing related representative discoveries in bone physiologies and pathologies. It also indicates current limitations, further refinement, and extended application of intravital imaging in bone research. Further details can be found in the article by Yuhao Liu, Quan Yuan, and Shiwen Zhanget ( e201960075 ).
Multivariate imaging techniques such as dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) have been shown to
provide valuable information for medical diagnosis. Even though these techniques provide new information, integrating and
evaluating the much wider range of information is a challenging task for the human observer. This task may be assisted with
the use of image fusion algorithms. 相似文献
To exploit the flood of data from advances in high throughput imaging of optically sectioned nuclei, image analysis methods
need to correctly detect thousands of nuclei, ideally in real time. Variability in nuclear appearance and undersampled volumetric
data make this a challenge. 相似文献
Magnetic resonance imaging (MRI) was used to study the hand and wrist in very early rheumatoid arthritis (RA), and the results
were compared with early and established disease. 相似文献