Renal-Clearable Organic Probes From D-A-D Type Aza-BODIPY Fluorophores for Multiphoton Deep-Brain Imaging.

Bright near-infrared (NIR) fluorescent probes play an important role in in vivo optical imaging. Here, renal-clearable nanodots prepared from Aza-BODIPY are reported fluorophores for multiphoton brain imaging. The design of donor-acceptor-donor (D-A-D) type conjugated structures endowed the fluorophores with large three-photon absorption cross-section for both 1620 and 2200 nm excitation. The side chain modification and lipid encapsulation yield ultrasmall nanodots (≈4 nm) and a high fluorescence quantum yield (≈0.35) at 720 nm emission in the aqueous phase. The measured three-photon action cross-section of a single Aza-BODIPY fluorophore in the nanodots is ≈30 times higher than the commonly used Sulforhodamine 101 dye. Three-photon deep brain imaging of subcortical structures is demonstrated, reaching a depth of 1900 µm below the brain surface in a live mouse study. The nanodots enabled blood flow measurement at a depth of 1617 µm using line scanning three-photon microscopy (3PM). This work provides superior fluorescent probes for multiphoton deep-brain imaging.

AutoLNMNet: Automated Network for Estimating Lymph-Node Metastasis in EGC Using a Pyramid Vision Transformer and Data Derived From Multiphoton Microscopy.

Lymph-node status is important in decision-making during early gastric cancer (EGC) treatment. Currently, endoscopic submucosal dissection is the mainstream treatment for EGC. However, it is challenging for even experienced endoscopists to accurately diagnose and treat EGC. Multiphoton microscopy can extract the morphological features of collagen fibers from tissues. The characteristics of collagen fibers can be used to assess the lymph-node metastasis status in patients with EGC. First, we compared the accuracy of four deep learning models (VGG16, ResNet34, MobileNetV2, and PVTv2) in training preprocessed images and test datasets. Next, we integrated the features of the best-performing model, which was PVTv2, with manual and clinical features to develop a novel model called AutoLNMNet. The prediction accuracy of AutoLNMNet for the no metastasis (Ly0) and metastasis in lymph nodes (Ly1) stages reached 0.92, which was 0.3% higher than that of PVTv2. The receiver operating characteristics of AutoLNMNet in quantifying Ly0 and Ly1 stages were 0.97 and 0.97, respectively. Therefore, AutoLNMNet is highly reliable and accurate in detecting lymph-node metastasis, providing an important tool for the early diagnosis and treatment of EGC.

Ultralow-dose irradiation enables engraftment and intravital tracking of disease initiating niches in clonal hematopoiesis.

Recent advances in imaging suggested that spatial organization of hematopoietic cells in their bone marrow microenvironment (niche) regulates cell expansion, governing progression, and leukemic transformation of hematological clonal disorders. However, our ability to interrogate the niche in pre-malignant conditions has been limited, as standard murine models of these diseases rely largely on transplantation of the mutant clones into conditioned mice where the marrow microenvironment is compromised. Here, we leveraged live-animal microscopy and ultralow dose whole body or focal irradiation to capture single cells and early expansion of benign/pre-malignant clones in the functionally preserved microenvironment. 0.5 Gy whole body irradiation (WBI) allowed steady engraftment of cells beyond 30 weeks compared to non-conditioned controls. In-vivo tracking and functional analyses of the microenvironment showed no change in vessel integrity, cell viability, and HSC-supportive functions of the stromal cells, suggesting minimal inflammation after the radiation insult. The approach enabled in vivo imaging of Tet2+/- and its healthy counterpart, showing preferential localization within a shared microenvironment while forming discrete micro-niches. Notably, stationary association with the niche only occurred in a subset of cells and would not be identified without live imaging. This strategy may be broadly applied to study clonal disorders in a spatial context.

Near-Infrared-II Photoactivated Iron(III) Complexes for Highly Efficient RNS and ROS Synergistic Therapy.

Reactive nitrogen species (RNS) are more lethal than reactive oxygen species (ROS), which gives them a very promising future in the field of cancer treatment. However, there are still a few drugs available for RNS generation. In this work, two 5th-order nonlinear optical materials, FB-Fe(III)/SNP@PEG and FB-Fe(II)-FB/SNP@PEG, are synthesized. The outstanding nonlinear optical properties of FB-Fe(III)/SNP@PEG help to achieve generation of bounteous superoxide anions (O2•-) in deep tissues, while sodium nitroprusside (SNP) provides NO in the body, both of which are prerequisites for RNS generation. Meanwhile, type I and type II ROS were also generated under irradiation of a 1600 nm laser. Based on the synergistic effect of ROS and RNS, FB-Fe(III)/SNP@PEG induced mitochondrial damage and DNA fragmentation and inhibited tumor cells through apoptosis, possessing better therapeutic effects than FB-Fe(II)-FB/SNP@PEG. This work put forward an innovative strategy to achieve the cooperative release of RNS and ROS in deep tissues, which provides insights and ideas for applying nonlinear optical materials to RNS therapy.

Fast Multiphoton Microscopic Imaging Joint Image Super-Resolution for Automated Gleason Grading of Prostate Cancers.

Gleason grading system is dependable for quantifying prostate cancer. This paper introduces a fast multiphoton microscopic imaging method via deep learning for automatic Gleason grading. Due to the contradiction between multiphoton microscopy (MPM) imaging speed and quality, a deep learning architecture (SwinIR) is used for image super-resolution to address this issue. The quality of low-resolution image is improved, which increased the acquisition speed from 7.55 s per frame to 0.24 s per frame. A classification network (Swin Transformer) was introduced for automated Gleason grading. The classification accuracy and Macro-F1 achieved by training on high-resolution images are respectively 90.9% and 90.9%. For training on super-resolution images, the classification accuracy and Macro-F1 are respectively 89.9% and 89.9%. It shows that super-resolution image can provide a comparable performance to high-resolution image. Our results suggested that MPM joint image super-resolution and automatic classification methods hold the potential to be a real-time clinical diagnostic tool for prostate cancer diagnosis.

Multitask machine learning-based tumor-associated collagen signatures predict peritoneal recurrence and disease-free survival in gastric cancer.

BACKGROUND: Accurate prediction of peritoneal recurrence for gastric cancer (GC) is crucial in clinic. The collagen alterations in tumor microenvironment affect the migration and treatment response of cancer cells. Herein, we proposed multitask machine learning-based tumor-associated collagen signatures (TACS), which are composed of quantitative collagen features derived from multiphoton imaging, to simultaneously predict peritoneal recurrence (TACSPR) and disease-free survival (TACSDFS). METHODS: Among 713 consecutive patients, with 275 in training cohort, 222 patients in internal validation cohort, and 216 patients in external validation cohort, we developed and validated a multitask machine learning model for simultaneously predicting peritoneal recurrence (TACSPR) and disease-free survival (TACSDFS). The accuracy of the model for prediction of peritoneal recurrence and prognosis as well as its association with adjuvant chemotherapy were evaluated. RESULTS: The TACSPR and TACSDFS were independently associated with peritoneal recurrence and disease-free survival in three cohorts, respectively (all P < 0.001). The TACSPR demonstrated a favorable performance for peritoneal recurrence in all three cohorts. In addition, the TACSDFS also showed a satisfactory accuracy for disease-free survival among included patients. For stage II and III diseases, adjuvant chemotherapy improved the survival of patients with low TACSPR and low TACSDFS, or high TACSPR and low TACSDFS, or low TACSPR and high TACSDFS, but had no impact on patients with high TACSPR and high TACSDFS. CONCLUSIONS: The multitask machine learning model allows accurate prediction of peritoneal recurrence and survival for GC and could distinguish patients who might benefit from adjuvant chemotherapy.

Dramatically Enhancing Multiphoton Harvesting Metal-Organic Frameworks for NIR-II Photocatalysis through Functional Regulation of Octupolar Molecules.

The development of effective multiphoton absorption (MPA) materials for near-infrared (NIR) light-driven photocatalysis holds great significance. In this study, we incorporated two multibranched cyclometallated iridium(III) modules with varying degrees of conjugation onto MPA-inert metal-organic frameworks (MOFs) to active MPA performance. Subsequently, the MOFs were further modified with Co(II) and hyaluronic acid (HA) to fabricate MINCH and MISCH, respectively. By introducing octupolar molecules and expanding the conjugation, MISCH exhibited a larger MPA cross section for efficient NIR light absorption and improved carrier transfer, leading to outstanding NIR light-driven multiphoton photocatalytic hydrogen production. Moreover, the HA modification enabled MISCH to achieve specific multiphoton photocatalytic hydrogen therapy for cancer cells. This study provides valuable insights into constructing highly active MPA materials for NIR light-driven photocatalysis, presenting a potential platform for hydrogen therapy in tumor treatment.

Manipulating Charge Distribution of Graphitic Carbon Nitride for Boosting NIR-II Light-Activated Reactive Oxygen Species Generation.

Structure engineering is of great importance to enhance the carrier separation efficiency of multiphoton absorption (MPA) materials for near-infrared (NIR) light-driven reactive oxygen species (ROS) generation. In this study, the MPA-responsive potassium/cyano group-functionalized graphitic carbon nitride was investigated, demonstrating charge redistribution and improved carrier separation efficiency by density functional theory calculations and experimental results. With various types of boosted ROS generation under UV-vis or NIR-II light irradiation, the potassium/cyano group-functionalized graphitic carbon nitride could achieve efficient multiphoton photodynamic therapy after reducing the particle size. This study developed a simple strategy to manipulate charge distribution for booting NIR light-activated ROS generation in efficient multiphoton photodynamic therapy.

Near-Infrared Emissive π-Conjugated Oligomer Nanoparticles for Three- and Four-Photon Deep-Brain Microscopic Imaging Beyond 1700 nm Excitation.

High-resolution visualization of the deep brain is still a challenging and very significant issue. Multiphoton microscopy (MPM) holds great promise for high-spatiotemporal deep-tissue imaging under NIR-III and NIR-IV excitation. However, thus far, their applications have been seriously restricted by the scarcity of efficient organic probes. Herein, we designed and synthesized two donor-acceptor-donor-type conjugated small molecules (TNT and TNS) for in vivo mouse deep-brain imaging with three- and four-photon microscopy under 1700 and 2200 nm excitation. With a selenium (Se) substitution, we synthesized two conjugated small molecules to promote their emission into the deep near-infrared region with high quantum yields of 55% and 20% in THF solvent, respectively, and their water-dispersive nanoparticles have relatively large absorption cross-sections in the 1700 and 2200 nm windows, respectively, with good biosafety. With these superiorities, these organic NPs achieve high-resolution deep-brain imaging via three-photon and four-photon microscopy with excitation at 1700 and 2200 nm windows, and 1620 µm deep in the brain vasculature can be visualized in vivo. This study demonstrates the efficiency of NIR-emissive conjugated small molecules for high-performance MPM imaging in the NIR-III and NIR-IV window and provides a route for the future design of organic MPM probes.

3D Printed Microneedles for the Transdermal Delivery of NAD+ Precursor: Toward Personalization of Skin Delivery.

3D printing of microneedles (µNDs) for transdermal therapy has the potential to enable patient personalization based on the target disease, site of application, and dosage requirements. To convert this concept to reality, it is necessary that the 3D printing technology can deliver high resolution, an affordable cost, and large print volumes. With the introduction of benchtop 4K and 8K 3D printers, it is now possible to manufacture medical devices like µNDs at sufficient resolution and low cost. In this research, we systematically optimized the 3D printing design parameters such as resin viscosity, print angle, layer height, and curing time to generate customizable µNDs. We have also developed an innovative 3D coating microtank device to optimize the coating method. We have applied this to the development of novel µNDs to deliver an established NAD+ precursor molecule, nicotinamide mononucleotide (NMN). A methacrylate-based polymer photoresin (eSun resin) was diluted with methanol to adjust the resin viscosity. The 3D print layer height of 25 µm yielded a smooth surface, thus reducing edge-ridge mismatches. Printing µNDs at 90° to the print platform yielded 84.28 ± 2.158% (n = 5) of the input height thus increasing the tip sharpness (48.52 ± 10.43 µm, n = 5). The formulation containing fluorescein (model molecule), sucrose (viscosity modifier), and Tween-20 (surface tension modifier) was coated on the µNDs using the custom designed microtank setup, and the amount deposited was determined fluorescently. The dye-coated µND arrays inserted into human skin (in vitro) showed a fluorescence signal at a depth of 150 µm (n = 3) into the skin. After optimization of the 3D printing parameters and coating protocol using fluorescein, NMN was coated onto the µNDs, and its diffusion was assessed in full-thickness human skin in vitro using a Franz diffusion setup. Approximately 189 ± 34.5 µg (5× dipped coated µNDs) of NMN permeated through the skin and 41.2 ± 7.53 µg was left in the skin after 24 h. Multiphoton microscopy imaging of NMN-coated µND treated mouse ear skin ex vivo demonstrated significantly (p < 0.05) increased free-unbound NADPH and reduced fluorescence lifetime of NADPH, both of which are indicative of cellular metabolic rates. Our study demonstrates that low-cost benchtop 3D printers can be used to print high-fidelity µNDs with the ability to rapidly coat and release NMN which consequently caused changes in intracellular NAD+ levels.

Harnessing Metal-Organic Frameworks for NIR-II Light-Driven Multiphoton Photocatalytic Water Splitting in Hydrogen Therapy.

The construction of near-infrared (NIR) light-activated hydrogen-producing materials that enable the controlled generation and high-concentration release of hydrogen molecules in deep tumor tissues and enhance the effects of hydrogen therapy holds significant scientific importance. To address the key technical challenge of low-efficiency oxidation-reduction reactions for narrow-bandgap photocatalytic materials, this work proposes an innovative approach for the controllable fabrication of multiphoton photocatalytic materials to overcome the limitations imposed by traditional near-infrared photocatalysts with "narrow-bandgap" constraints. Herein, an NIR-responsive multiphoton photocatalyst, ZrTc-Co, is developed by utilizing a post-synthetic coordination modification strategy to introduce hydrogenation active site CoII into a multiphoton responsive MOF (ZrTc). The results reveal that with the introduction of the CoII site, electron-hole recombination can be efficiently suppressed, thus promoting the efficiency of hydrogen evolution reaction. In addition, the integration of CoII can effectively enhance charge transfer and improve static hyperpolarizability, which endows ZrTc-Co with excellent multiphoton absorption. Moreover, hyaluronic acid modification endows ZrTc-Co with cancer cell-specific targeting characteristics, laying the foundation for tumor-specific elimination. Collectively, the proposed findings present a strategy for constructing NIR-II light-mediated hydrogen therapeutic agents for deep tumor elimination.

Intradermal Delivery of Calcium Hydroxylapatite With Fractionated Ablation.

OBJECTIVES: The absorption of biostimulatory particulate matter following its application to fractional skin defects remains poorly understood, and even less is known about its in vivo impact in terms of tissue integration. The objectives of this study are twofold: (1) to evaluate the potential of calcium hydroxylapatite (CaHA) to penetrate through skin treated with a fractional laser; and (2) to assess the effectiveness of clinical laser scanning microscopy technologies in monitoring the effects of such treatment over time. METHODS: One area on a volunteer's arm was treated with a fractional erbium laser (Sciton Inc., Palo Alto, CA), while a second area received the same laser treatment followed by CaHA topical application. We used reflectance confocal microscopy (RCM) and multiphoton microscopy (MPM) to noninvasively image beneath the surface of the treated skin to study and monitor the effects of these treatments within 1 h of treatment and at four additional time points over a 6-week period. RESULTS: One hour posttreatment, at different depths beneath the skin surface, MPM and RCM provided similar visualizations of laser-induced channels. In skin treated by both laser and CaHA, these two imaging methods provided complementary information. RCM captured the lateral and depth distribution of CaHA microspheres and were seen as bright spheres as they became incorporated into the healing tissue. MPM, meanwhile, visualized the CaHA microparticles as dark shadow spheres within the laser-induced channels and encroaching healing tissue. Furthermore, MPM provided critical information about collagen regeneration around the microspheres, with the collagen visually marked by its distinct second harmonic generation (SHG) signal. CONCLUSIONS: This observational pilot study demonstrates that CaHA, a collagen stimulator used as a dermal filler, can not only be inserted into the dermis after fractional laser treatment but remains in the healing skin for at least 6 weeks posttreatment. The noninvasive imaging techniques RCM and MPM successfully captured the presence of CaHA microspheres mid-dermis during the healing phase. They also demonstrated new collagen production around the microspheres, highlighting the effectiveness of these imaging approaches in monitoring such treatment over time.

Perovskites as Multiphoton Fluorescence Contrast Agents for In Vivo Imaging.

Multiphoton fluorescence microscopy is a powerful tool for imaging and exploring biological tissue at subcellular spatial resolution while minimizing photobleaching and autofluorescence. For optimal performance in multiphoton microscopy, materials exhibiting a large multiphoton absorption cross section (σn) and fluorescence quantum yield are desired. Notably, perovskite nanocrystals (CsPbX3, PNCs) exhibit exceptionally large two-, three-, up to five photon absorption cross section (σ2 ∼ 106 GM, σ3 ∼ 10-73 cm6s2 photon-2, σ5 ∼ 10-136 cm10s4 photon-4), along with near unity fluorescence quantum yield, making them desirable for deep tissue applications. Here, we employed PNCs as contrast agents to image mesenchymal stromal cells in a living mouse. The PNCs were stabilized by encapsulating them in a SiO2 matrix (∼60-70 nm in diameter), offering versatility for subsequent surface modification to target specific biological entities for both diagnostic and therapeutic applications. Multiphoton imaging of PNCs offers substantial benefits for dynamic tracking of cells in deep tissue, such as in understanding immune cell migration and other biological processes in both healthy and diseased tissues.

Bone turnover markers in the preoperative assessment of bone quality - A prospective investigation of bone microstructure and advanced glycation endproducts in lumbar fusion patients.

INTRODUCTION: Effective tools to evaluate bone quality preoperatively are scarce and the standard method to determine bone quality requires an invasive biopsy. A non-invasive, and preoperatively available method for bone quality assessment would be of clinical value. The purpose of this study is to investigate the associations of bone formation marker, serum bone alkaline phosphatase (BAP), and bone resorption marker, urine collagen cross-linked N-telopeptide (uNTX) to volumetric bone mineral density (vBMD), fluorescent advanced glycation endproducts (fAGEs) and bone microstructure. MATERIALS AND METHODS: A cross-secional analysis using prospective data of patients undergoing lumbar spinal fusion was performed. BAP and uNTX were preoperatively collected. Quantitative computed tomography (QCT) was performed at the lumbar spine (vBMD ≤ 120 mg/cm3 osteopenic/osteoporotic). Bone biopsies from the posterior superior iliac spine were obtained and evaluated with multiphoton fluorescence microscopy for fAGEs and microcomputed tomography (µCT) for bone microarchitecture. Correlations between BAP/uNTX to vBMD, fAGEs and µCT parameters were assessed with Spearman's ρ. Receiver operating characteristic (ROC) analysis evaluated BAP and uNTX as predictors for osteopenia/osteoporosis. Multivariable linear regression models adjusting for age, sex, BMI, race and diabetes mellitus determined associations between BAP/uNTX and fAGEs. RESULTS: 127 prospectively enrolled patients (50.4% female, 62.5 years, BMI 28.7 kg/m2) were analyzed. uNTX (ρ=-0.331,p < 0.005) and BAP (ρ=-0.245,p < 0.025) decreased with cortical fAGEs, and uNTX (ρ=-0.380,p < 0.001) decreased with trabecular fAGEs. BAP and uNTX revealed no significant correlation with vBMD. ROC analysis for BAP and uNTX discriminated osteopenia/osteoporosis with AUC of 0.477 and 0.561, respectively. In the multivariable analysis, uNTX decreased with increasing trabecular fAGEs after adjusting for covariates (ß = 0.923;p = 0.031). CONCLUSION: This study demonstrated an inverse association of bone turnover markers and fAGEs. Both uNTX and BAP could not predict osteopenia/osteoporosis in the spine. uNTX reflects collagen characteristics and might have a complementary role to vBMD, as a non-invasive tool for bone quality assessment in spine surgery.

Using Multiphoton Intravital Microscopy to Study Neutrophil Transmigration and Blood-Brain Barrier Permeability in a Mouse Model of Herpes Simplex Virus Encephalitis.

Multiphoton intravital microscopy (MP-IVM) is an imaging technique used for the observation of living organisms at a microscopic resolution. The tissue of interest is exposed through a window allowing imaging of cells in real time. Using MP-IVM, the temporospatial kinetics of leukocyte transendothelial migration can be visualized and quantitated using reporter mice and cell-specific fluorophore-conjugated monoclonal antibodies to track the leukocytes within and outside of vascular beds. Here we describe a method used to study neutrophil transendothelial migration and blood-brain barrier permeability in a mouse model of herpes simplex virus I (HSV) encephalitis.

Multiphoton excited polymerized biomimetic models of collagen fiber morphology to study single cell and collective migration dynamics in pancreatic cancer.

The respective roles of aligned collagen fiber morphology found in the extracellular matrix (ECM) of pancreatic cancer patients and cellular migration dynamics have been gaining attention because of their connection with increased aggressive phenotypes and poor prognosis. To better understand how collagen fiber morphology influences cell-matrix interactions associated with metastasis, we used Second Harmonic Generation (SHG) images from patient biopsies with Pancreatic ductal adenocarcinoma (PDAC) as models to fabricate collagen scaffolds to investigate processes associated with motility. Using the PDAC BxPC-3 metastatic cell line, we investigated single and collective cell dynamics on scaffolds of varying collagen alignment. Collective or clustered cells grown on the scaffolds with the highest collagen fiber alignment had increased E-cadherin expression and larger focal adhesion sites compared to single cells, consistent with metastatic behavior. Analysis of single cell motility revealed that the dynamics were characterized by random walk on all substrates. However, examining collective motility over different time points showed that the migration was super-diffusive and enhanced on highly aligned fibers, whereas it was hindered and sub-diffusive on un-patterned substrates. This was further supported by the more elongated morphology observed in collectively migrating cells on aligned collagen fibers. Overall, this approach allows the decoupling of single and collective cell behavior as a function of collagen alignment and shows the relative importance of collective cell behavior as well as fiber morphology in PDAC metastasis. We suggest these scaffolds can be used for further investigations of PDAC cell biology. STATEMENT OF SIGNIFICANCE: Pancreatic ductal adenocarcinoma (PDAC) has a high mortality rate, where aligned collagen has been associated with poor prognosis. Biomimetic models representing this architecture are needed to understand complex cellular interactions. The SHG image-based models based on stromal collagen from human biopsies afford the measurements of cell morphology, cadherin and focal adhesion expression as well as detailed motility dynamics. Using a metastatic cell line, we decoupled the roles of single cell and collective cell behavior as well as that arising from aligned collagen. Our data suggests that metastatic characteristics are enhanced by increased collagen alignment and that collective cell behavior is more relevant to metastatic processes. These scaffolds provide new insight in this disease and can be a platform for further experiments such as testing drug efficacy.

A three-dimensional valve-on-chip microphysiological system implicates cell cycle progression, cholesterol metabolism and protein homeostasis in early calcific aortic valve disease progression.

BACKGROUND: Calcific aortic valve disease (CAVD) is one of the most common forms of valvulopathy, with a 50 % elevated risk of a fatal cardiovascular event, and greater than 15,000 annual deaths in North America alone. The treatment standard is valve replacement as early diagnostic, mitigation, and drug strategies remain underdeveloped. The development of early diagnostic and therapeutic strategies requires the fabrication of effective in vitro valve mimetic models to elucidate early CAVD mechanisms. METHODS: In this study, we developed a multilayered physiologically relevant 3D valve-on-chip (VOC) system that incorporated aortic valve mimetic extracellular matrix (ECM), porcine aortic valve interstitial cell (VIC) and endothelial cell (VEC) co-culture and dynamic mechanical stimuli. Collagen and glycosaminoglycan (GAG) based hydrogels were assembled in a bilayer to mimic healthy or diseased compositions of the native fibrosa and spongiosa. Multiphoton imaging and proteomic analysis of healthy and diseased VOCs were performed. RESULTS: Collagen-based bilayered hydrogel maintained the phenotype of the VICs. Proteins related to cellular processes like cell cycle progression, cholesterol biosynthesis, and protein homeostasis were found to be significantly altered and correlated with changes in cell metabolism in diseased VOCs. This study suggested that diseased VOCs may represent an early, adaptive disease initiation stage, which was corroborated by human aortic valve proteomic assessment. CONCLUSIONS: In this study, we developed a collagen-based bilayered hydrogel to mimic healthy or diseased compositions of the native fibrosa and spongiosa layers. When the gels were assembled in a VOC with VECs and VICs, the diseased VOCs revealed key insights about the CAVD initiation process. STATEMENT OF SIGNIFICANCE: Calcific aortic valve disease (CAVD) elevates the risk of death due to cardiovascular pathophysiology by 50 %, however, prevention and mitigation strategies are lacking, clinically. Developing tools to assess early disease would significantly aid in the prevention of disease and in the development of therapeutics. Previously, studies have utilized collagen and glycosaminoglycan-based hydrogels for valve cell co-cultures, valve cell co-cultures in dynamic environments, and inorganic polymer-based multilayered hydrogels; however, these approaches have not been combined to make a physiologically relevant model for CAVD studies. We fabricated a bi-layered hydrogel that closely mimics the aortic valve and used it for valve cell co-culture in a dynamic platform to gain mechanistic insights into the CAVD initiation process using proteomic and multiphoton imaging assessment.

In vivo organoid growth monitoring by stimulated Raman histology.

Patient-derived tumor organoids have emerged as a crucial tool for assessing the efficacy of chemotherapy and conducting preclinical drug screenings. However, the conventional histological investigation of these organoids necessitates their devitalization through fixation and slicing, limiting their utility to a single-time analysis. Here, we use stimulated Raman histology (SRH) to demonstrate non-destructive, label-free virtual staining of 3D organoids, while preserving their viability and growth. This novel approach provides contrast similar to conventional staining methods, allowing for the continuous monitoring of organoids over time. Our results demonstrate that SRH transforms organoids from one-time use products into repeatable models, facilitating the efficient selection of effective drug combinations. This advancement holds promise for personalized cancer treatment, allowing for the dynamic assessment and optimization of chemotherapy treatments in patient-specific contexts.

Accessing the Low-Polar Molecular Composition of Boreal and Arctic Peat-Burning Organic Aerosol via Thermal Analysis and Ultrahigh-Resolution Mass Spectrometry: Structural Motifs and Their Formation.

Peatland fires emit organic carbon-rich particulate matter into the atmosphere. Boreal and Arctic peatlands are becoming more vulnerable to wildfires, resulting in a need for better understanding of the emissions of these special fires. Extractable, nonpolar, and low-polar organic aerosol species emitted from laboratory-based boreal and Arctic peat-burning experiments are analyzed by direct-infusion atmospheric pressure photoionization (APPI) ultrahigh-resolution mass spectrometry (UHRMS) and compared to time-resolved APPI UHRMS evolved gas analysis from the thermal analysis of peat under inert nitrogen (pyrolysis) and oxidative atmosphere. The chemical composition is characterized on a molecular level, revealing abundant aromatic compounds that partially contain oxygen, nitrogen, or sulfur and are formed at characteristic temperatures. Two main structural motifs are identified, single core and multicore, and their temperature-dependent formation is assigned to the thermal degradation of the lignocellulose building blocks and other parts of peat.

Ultrafast Laser 3D Nanolithography of Fiber-Integrated Silica Microdevices.

Fiber-integrated micro/nanostructures play a crucial role in modern industry, mainly owing to their compact size, high sensitivity, and resistance to electromagnetic interference. However, the three-dimensional manufacturing of fiber-tip functional structures beyond organic polymers remains challenging. It is essential to construct fiber-integrated inorganic silica with designed functional nanostructures for microsystem applications. Here, we develop a strategy for the 3D nanolithography of fiber-integrated silica from hybrid organic-inorganic materials by ultrafast laser-induced multiphoton absorption. Without silica nanoparticles and polymer additives, the acrylate-functionalized precursors can be locally cross-linked through a nonlinear effect. Followed by annealing at low temperature, the as-printed micro/nanostructures are transformed to high-quality silica with sub-100 nm resolution. Silica microcantilever probes and microtoroid resonators are directly integrated onto the optical fiber, showing strong thermal stability and quality factors. This work provides a promising strategy for fabricating desired fiber-tip silica micro/nanostructures, which is helpful for the development of integrated functional device applications.