At the present, the two clinical biomarkers used to monitor diabetic progression are blood glucose and HbA1c. However, advanced glycation end products (AGEs) have been shown to contribute to diabetic pathogenesis, and there is interest in the use of AGEs in tissues as long-term glycemic markers. In this study, we investigate the in vitro rate of fluorescent AGEs (fAGEs) formation with multiphoton microscopy in different porcine tissues (aorta, cornea, kidney, dermis, and tendon) from glucose, galactose, and fructose, three primary monosaccharides found in human diets. These results may be of value in developing long-term glycemic biomarkers for diabetes.
This work illustrates how tumor proportional score is estimated using object detection method YOLO and compared with a pathologist's calculation. Results show deep learning can achieve good results and be used on clinical applications.
At the present, identifying head and neck squamous cell carcinoma (HNSCC) patients for immune checkpoint inhibitor therapy (ICIT) is achieved through the determination of Tumor Proportion Score (TPS) or the percentage of tumor cells positively labeled for PD-L1. Estimation of TPS is largely done in a manual fashion by a trained pathologist. In the case of HNSCC, the histological section can be over 1 cm in size in which over 100,000 cancer cells need to be evaluated for PD-L1 expression. To expedite the TPS evaluation process for such large specimens, we have developed a platform in which artificial intelligence (AI) is used for TPS determination. With additional development, this approach may be used in the clinical setting to assist pathologists in TPS evaluation.
Most of the cornea is organized through a lattice pattern of collagen fibrils which is responsible for its transparency. Developing avian cornea is a highly organized extracellular matrix composed largely of striated collagen fibrils. In this study, we use embryonic corneas from chicks as model animal to study the temporal and spatial variation of corneal stroma. Through the use of Fast Fourier Transform second harmonic generation microscopy, we quantified collagen alignment of the entire corneal thickness during development. Corneal structural variation at different stage of developing embryos was studied. These results indicate that both the rotational pitch and overall rotational angle of corneal stroma is highly involved in the developing process of cornea, and these properties exhibit strong correlation during the development of left and right corneas.
In nonlinear optical imaging of biological samples, epi-illumination collection of fluorescent signals results in half of the signal loss. We enhanced the collected luminescent signal by using a multilayer coating of tantalum pentoxide (Ta2O5) and cerium oxide (SiO2). Our coatings are biocompatible, allowing visual inspection of samples and optimizing the collection of luminescent signals. This method was confirmed on a number of samples, including sulforhodamine solutions, fluorescent microspheres and labeled 3T3 cells. In all cases, the coated coverslips were used to enhance the signal by approximately 2 times. Image analysis also shows that signal enhancement does not result in additional photobleaching. Our results indicate that the designed coated coverslip enhances multi-photon microscopy detection signals.
Second order susceptibility microscopy was used to image and characterize chondrogenesis in cultured human mesesnchymal stem cells (hMSCs). Second order susceptibility analysis shows that the second order susceptibility tensor ratios can be used to characterize type I and II collagens in living tissues and that both collagen types are produce at the onset of chondrogenesis. Time-lapse analysis shows that relative to type I collagen, type II collagen increases with time. Eventually, type II collagen content stabilizes at the composition of 70% of total collagen content. This approach can be used to continuously and non-invasively monitor the production of collagens I/II and developed into an effective quality control tool for monitoring extracellular matrix production in engineered tissues.
We used intravital multiphoton microscopy to study the recovery of hepatobiliary metabolism following carbon tetrachloride (CCl4) induced hepatotoxicity in mouse. Our images were processed by a first order kinetic model to generate rate constant resolved images of the mouse liver. At Day 14 following induction, a restoration of the mouse hepatobiliary function was found. Our approach allows the study of the response of hepatic functions to chemical agents in real time and is useful for studying pharmacokinetics of drug molecules through optical microscopic imaging.
The cornea is a transparent tissue with significant refractive and barrier functions. The adult cornea has five layers: an outer epithelium layer, a Bowman’s layer which forms the transition between the corneal epithelium and the underlying stroma, a middle stromal layer of collagen-rich extracellular matrix between stromal keratocytes, a Descemet's membrane which separates the stroma from the underlying endothelial layer of the cornea and an inner layer of endothelial cells. Early studies showed that the that collagen fibrils of chick cornea display orthogonal-like pattern. Moreover, primary and secondary stroma have been identified in developing cornea. The primary stroma of the developing avian cornea is a highly organized extracellular matrix composed largely of striated collagen fibrils synthesized by the epithelium. Previous studies have found at least two different fibrillar collagen types, such as type I and II, are shown during the development. Other type of collagen, such as type IX, also appears to be involved in the collagen assembly process. This matrix is subsequently invaded by periocular mesenchymal cells which secrete a secondary cornea stroma to gradually replace the primary one. This indicates that the development of embryonic cornea involves many important biological events and exhibits highly dynamic characteristics. Since second-harmonic generation (SHG) is a nonlinear second order optical process which occurs in noncentrosymmetric systems with a large hyperpolarizability, it has emerged as a powerful modality for imaging fibrillar collagen in a diverse range of tissues. It is highly sensitive to the collagen fibril/fiber structure, and to changes that occur in diseases such as cancer, fibrosis and connective tissue disorders. To explore the structural variation of embryonic cornea, we use Fast Fourier Transform second harmonic generation microscopy as a tool in our study.
Intravital multiphoton microscopy was used to study hepatobiliary metabolism in chronic pathologies of the liver. Through the use of the probe molecule 6-carboxyfluorescein diacetate (6-CFDA), the effects of liver fibrosis, fatty liver, and hepatocellular carcinoma on the metabolic capabilities of mouse liver were investigated. After the acquisition of time lapse images, a first order kinetic model was used to calculate rate constant resolved images of various pathologies. It was found the ability of the liver to metabolically process the probe molecules varies among different pathologies, with liver fibrosis and fatty liver disease negatively impacted the uptake, processing, and excretion of molecules.
Cornea is one of the collagen-rich connective tissues and plays an important role in vision. While X-ray scattering techniques are able to determine bulk structure of cornea, second harmonic generation microscopy can reveal depthdependent details of the corneal stroma comprised of a layered network of fibrillar collagen. In this work, we used Fast Fourier Transform second harmonic generation microscopy as a tool to determine the directionality of corneal stroma as a function of depth. Our results also display the position dependent difference of corneal stroma architecture.
Second order susceptibility (SOS) microscopy is used as a contrast mechanism for distinguishing types I and III collagen in tissues. In addition, relative proportion of collagen type was determined by analyzing the histograms of SOS ratios. Our results show that second order susceptibility is an effective metric for differentiating types I and III collagen without extrinsic labeling.
We used intravital multiphoton microscopy to study the recovery of hepatobiliary metabolism following carbon tetrachloride (CCl4) induced hepatotoxicity in mice. The acquired images were processed by a first order kinetic model to generate rate constant resolved images of the mouse liver. We found that with progression of hepatotoxicity, the spatial gradient of hepatic function disappeared. A CCl4-induced damage mechanism involves the compromise of membrane functions, resulting in accumulation of processed 6-carboxyfluorescein molecules. At day 14 following induction, a restoration of the mouse hepatobiliary function was found. Our approach allows the study of the response of hepatic functions to chemical agents in real time and is useful for studying pharmacokinetics of drug molecules through optical microscopic imaging.
The unique organization of the corneal stromal collagen is responsible for mechanical strength and optical clarity of the
eye. However, factors and reasons on formation of the corneal stroma is still not fully understood. Second-harmonic
generation (SHG) is a nonlinear second order optical process occurring in noncentrosymmetric systems with a large
hyperpolarizability. Through the combination of the second harmonic generation (SHG) microcopy and optimized
Fourier-transform analysis, mature chicken corneas are investigated to probe the depth-dependent collagen organization
of the corneal stroma. Our results show that the anterior stroma behaves like a fan-like distribution of successively and
counterclockwisely rotated fibrous lamellae for paired corneas from the same chicken. However, the posterior stroma
maintains a non-rotating pattern while increasing in depth. Surprisingly, the thickness of the anterior stroma remains
almost constant throughout the temporal-nasal direction, but the posterior stroma does not behave the same. Through
quantitative analysis, the natural transition of the anterior and posterior stroma is also determined. These findings
enhance our understanding of the collagen-rich tissue in the chicken cornea model. Moreover, the Fourier-transformbased
modality, in combination with SHG microscopy, serves as a promising tool to determine collagen alignment in
embryonic development, tissue engineering and corneal diseases.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.