Tissue Model Research Articles

Sensitive Cancer Hypoxia Detection via a Dual-Locking Fluorescence Response System Using Two Hypoxia Indicators.

Hypoxia is intricately associated with various diseases, including ischemia, vascular disorders, and cancer. Particularly in cancer cells, hypoxia promotes tumor growth, cell proliferation, migration, and invasion and enhances treatment resistance, making its detection crucial for cancer diagnosis and therapy. However, methods for detecting hypoxia are limited, often relying on single-detection systems. In this study, we developed a dual-lock-based fluorescent probe that selectively exhibits strong green fluorescence under hypoxic conditions due to simultaneous activity of nitroreductases (NTRs) and hydrogen sulfide (H2S), with a high signal-to-background ratio. The biocompatibility and photophysical properties of the probes were thoroughly investigated through both extracellular and intracellular experimental analyses. Among the synthesized naphthalimide-based probes, the dual-detection probe DNNC demonstrated excellent selectivity and sensitivity to the simultaneous activity of NTR/H2S compared to other single-detection probes. The performance of DNNC was applied to various organ-derived cancer cells and tumor tissue models such as HeLa cell sparoids, enabling spatiotemporal confocal fluorescence imaging and quantitative analysis of hypoxic levels in cancer. Our development of DNNC is expected to significantly advance cancer diagnosis and treatment by molecularly detecting hypoxia associated with cancer aggressiveness, therapy resistance, and unfavorable prognosis.

Determining Rates of Molecular Secretion from Supernatant Concentration Measurements in a 3D-Bioprinted Human Liver Tissue Model.

The secretion rate of albumin is a key indicator of function in liver tissue models used for hepatotoxicity and pharmacokinetic testing. However, it is not generally clear how to determine molecular secretion rates from measurements of the molecular concentration in supernatant media. Here, we develop computational and analytical models of molecular transport in an experimental system that enable determination of albumin secretion rates based on measurements of albumin concentration in supernatant media. The experimental system is a 3D-bioprinted human liver tissue construct embedded in a 3D culture environment made from packed microgel particles swollen in liquid growth media. The mathematical models reveal that the range of albumin synthesis rates necessary to match experimentally measured albumin concentrations corresponds to reaction-limited conditions, where a steady state of albumin spatial distribution is rapidly reached between media exchanges. Our results show that temporally resolved synthesis rates can be inferred from serial concentration measurements of collected supernatant media. This link is critical to confidently assessing in vitro tissue performance in applications where critical quality attributes must be quantified, like in drug development and screening.

Comparing continuum and direct fiber models of soft tissues. An ocular biomechanics example reveals that continuum models may artificially disrupt the strains at both the tissue and fiber levels.

Collagen fibers are the main load-bearing component of soft tissues but difficult to incorporate into models. Whilst simplified homogenization models suffice for some applications, a thorough mechanistic understanding requires accurate prediction of fiber behavior, including both detailed fiber-level strains and long-distance transmission. Our goal was to compare the performance of a continuum model of the optic nerve head (ONH) built using conventional techniques with a fiber model we recently introduced which explicitly incorporates the complex 3D organization and interaction of collagen fiber bundles [1]. To ensure a fair comparison, we constructed the continuum model with identical geometrical, structural, and boundary specifications as for the fiber model. We found that: 1) although both models accurately matched the intraocular pressure (IOP)-induced globally averaged displacement responses observed in experiments, they diverged significantly in their ability to replicate specific 3D tissue-level strain patterns. Notably, the fiber model faithfully replicated the experimentally observed depth-dependent variability of radial strain, the ring-like pattern of meridional strain, and the radial pattern of circumferential strain, whereas the continuum model failed to do so; 2) the continuum model disrupted the strain transmission along each fiber, a feature captured well by the fiber model. These results demonstrate limitations of the conventional continuum models that rely on homogenization and affine deformation assumptions, which render them incapable of capturing some complex tissue-level and fiber-level deformations. Our results show that the strengths of explicit fiber modeling help capture intricate ONH biomechanics. They potentially also help modeling other fibrous tissues.

Optimizing miRNA transfection for screening in precision cut lung slices.

Precision cut lung slices(PCLS) are complex 3D lung tissue models, which preserve the native microenvironment, including cell diversity and cell-matrix interactions. They are an innovative ex vivo platform that allows studying disease as well as the effects of therapeutic agents or regulatory molecules(e.g. miRNA). The aim of our study was to develop a protocol to transfect PCLS with miRNA using lipid nanoparticles (LNPs) to enable higher throughput screening of miRNA, obviating the need for custom stabilization and internalization approaches. 4mm diameter PCLS were generated using agarose-filled rodent lungs and a vibratome. TYE665 labelled scrambled miRNA was used to evaluate transfection efficacy of six different commercially available LNPs. Transfection efficacy was visualised using live high content fluorescence microscopy, followed by higher resolution confocal fluorescence microscopy in fixed PCLS. Metabolic activity and cellular damage were assessed using WST-1 and lactate dehydrogenase(LDH) release. Using a live staining kit containing a cell membrane impermeant nuclear dye, RedDot2TM, we established that cellular membranes in PCLS are permeable in the initial 24 hours of slicing but diminished thereafter. Therefore, all transfection experiments occurred at least 24 hours after slicing. All six commercially available LNPs enabled transfection without inducing significant cytotoxicity or impaired metabolic function. However, RNAiMAX and INTERFERin led to increases in transfection efficacy as compared to other LNPs, with detection possible as low as 25nM. Therefore, LNP-based transfection of miRNA is possible and can be visualized in live or fixed PCLS, enabling future higher throughput studies using diverse miRNAs.

A Mesenteric Fat-Derived Radiomic Model to Identify Colonic Fibrosis and Predict Treatment Response to Biologics in Chronic Ulcerative Colitis.

Evidence suggested the lesion of ulcerative colitis stretches beyond mucosa. The application of radiomics on ulcerative colitis fibrosis is unclear. We aimed to characterize the colonic fibrosis and treatment response to biologics in chronic ulcerative colitis using radiomic features extracted from bowel wall and mesenteric adipose tissue. Retrospective analysis of prospective database. This study was conducted in a single tertiary center. A total of 72 patients who underwent proctocolectomy and 47 patients who received biologics induction were included. Computed Tomography images were collected and radiomic features were extracted to develop radiomic models using logistic regression. Main outcome was colonic fibrosis, which was classified into mild and severe based on histological scoring. The area under curve of the bowel wall model to predict severe fibrosis was 0.931 (p < 0.001) and 0.869 (p < 0.001) in the training and test cohort, respectively. For mesenteric adipose tissue model, area under curve was 0.947 (p < 0.001) and 0.837 (p < 0.001), respectively. The mesenteric adipose tissue model was superior to bowel wall model (area under curve, 0.809, p < 0.001 and 0.722, p = 0.006) in predicting response to biologics in chronic ulcerative colitis. Retrospective single center study. Two radiomic models derived from bowel wall and mesenteric adipose tissue features readily predicted colonic fibrosis and treatment response of biologics in chronic ulcerative colitis. The mesentery harbors critical information and was essentially involved in fibrogenesis. See Video Abstract.

A point-of-research decision in synovial tissue engineering: Mesenchymal stromal cells, tissue derived fibroblast or CTGF-mediated mesenchymal-to-fibroblast transition

Rheumatoid arthritis (RA) and osteoarthritis (OA) are prevalent inflammatory joint diseases characterized by synovitis, cartilage, and bone destruction. Fibroblast-like synoviocytes (FLSs) of the synovial membrane are a decisive factor in arthritis, making them a target for future therapies. Developing novel strategies targeting FLSs requires advanced in vitro joint models that accurately replicate non-diseased joint tissue. This study aims to identify a cell source reflecting physiological synovial fibroblasts. Therefore, we newly compared the phenotype and metabolism of “healthy” knee-derived FLSs from patients with ligament injuries (trauma-FLSs) to mesenchymal stromal cells (MSCs), their native precursors. We differentiated MSCs into fibroblasts using connective tissue growth factor (CTGF) and compared selected protein and gene expression patterns to those obtained from trauma-FLSs and OA-FLSs. Based on these findings, we explored the potential of an MSC-derived synovial tissue model to simulate a chronic inflammatory response akin to that seen in arthritis. We have identified MSCs as a suitable cell source for synovial tissue engineering because, despite metabolic differences, they closely resemble human trauma-derived FLSs. CTGF-mediated differentiation of MSCs increased HAS2 expression, essential for hyaluronan synthesis. It showed protein expression patterns akin to OA-FLSs, including markers of ECM components and fibrosis, and enzymes leading to a shift in metabolism towards increased fatty acid oxidation. In general, cytokine stimulation of MSCs in a synovial tissue model induced pro-inflammatory and pro-angiogenic gene expression, hyperproliferation, and increased glucose consumption, reflecting cellular response in human arthritis. We conclude that MSCs can serve as a proxy to study physiological synovial processes and inflammatory responses. In addition, CTGF-mediated mesenchymal-to-fibroblast transition resembles OA-FLSs. Thus, we emphasize MSCs as a valuable cell source for tools in preclinical drug screening and their application in tissue engineering.

Microstructure of the cerebellum and its afferent pathways underpins dystonia in myoclonus dystonia.

Myoclonus dystonia due to a pathogenic variant in SGCE (MYC/DYT-SGCE) is a rare condition involving a motor phenotype associating myoclonus and dystonia. Dysfunction within the networks relying on the cortex, cerebellum, and basal ganglia was presumed to underpin the clinical manifestations. However, the microarchitectural abnormalities within these structures and related pathways are unknown. Here, we investigated the microarchitectural brain abnormalities related to the motor phenotype in MYC/DYT-SGCE. We used neurite orientation dispersion and density imaging, a multicompartment tissue model of diffusion neuroimaging, to compare microarchitectural neurite organization in MYC/DYT-SGCE patients and healthy volunteers (HVs). Neurite density index (NDI), orientation dispersion index (ODI), and isotropic volume fraction (ISOVF) were derived and correlated with the severity of motor symptoms. Fractional anisotropy (FA) and mean diffusivity (MD) derived from the diffusion tensor approach were also analyzed. In addition, we studied the pathways that correlated with motor symptom severity using tractography analysis. Eighteen MYC/DYT-SGCE patients and 24 HVs were analyzed. MYC/DYT-SGCE patients showed an increase of ODI and a decrease of FA within their motor cerebellum. More severe dystonia was associated with lower ODI and NDI and higher FA within motor cerebellar cortex, as well as with lower NDI and higher ISOVF and MD within the corticopontocerebellar and spinocerebellar pathways. No association was found between myoclonus severity and diffusion parameters. In MYC/DYT-SGCE, we found microstructural reorganization of the motor cerebellum. Structural change in the cerebellar afferent pathways that relay inputs from the spinal cord and the cerebral cortex were specifically associated with the severity of dystonia.

An antifibrotic compound that ameliorates hyperglycaemia and fat accumulation in cell and HFD mouse models.

Appropriate management of blood glucose levels and the prevention of complications are important in the treatment of diabetes. We have previously reported on a compound named HPH-15 that is not only antifibrotic but also AMP-activated protein kinase (AMPK)-activating. In this study, we evaluated whether HPH-15 is useful as a therapeutic medication for diabetes. We examined the effects of HPH-15 on AMPK activation, glucose uptake, fat accumulation and lactic acid production in L6-GLUT4, HepG2 and 3T3-L1 cells, as a model of muscle, liver and fat tissue, respectively. Additionally, we investigated the glucose-lowering, fat-accumulation-suppressing, antifibrotic and AMPK-activating effect of HPH-15 in mice fed a high-fat diet (HFD). HPH-15 at a concentration of 10 µmol/l increased AMPK activation, glucose uptake and membrane translocation of GLUT4 in each cell model to the same extent as metformin at 2 mmol/l. The production of lactic acid (which causes lactic acidosis) in HPH-15-treated cells was equal to or less than that observed in metformin-treated cells. In HFD-fed mice, HPH-15 lowered blood glucose from 11.1±0.3 mmol/l to 8.2±0.4 mmol/l (10 mg/kg) and 7.9±0.4 mmol/l (100 mg/kg) and improved insulin resistance. The HPH-15 (10 mg/kg) group showed the same level of AMPK activation as the metformin (300 mg/kg) group in all organs. The HPH-15-treated HFD-fed mice also showed suppression of fat accumulation and fibrosis in the liver and fat tissue; these effects were more significant than those obtained with metformin. Mice treated with high doses of HPH-15 also exhibited a 44% reduction in subcutaneous fat. HPH-15 activated AMPK at lower concentrations than metformin in vitro and in vivo and improved blood glucose levels and insulin resistance in vivo. In addition, HPH-15 was more effective than metformin at ameliorating fatty liver and adipocyte hypertrophy in HFD-fed mice. HPH-15 could be effective in preventing fatty liver, a common complication in diabetic individuals. Additionally, in contrast to metformin, high doses of HPH-15 reduced subcutaneous fat in HFD-fed mice. Presumably, HPH-15 has a stronger inhibitory effect on fat accumulation and fibrosis than metformin, accounting for the reduction of subcutaneous fat. Therefore, HPH-15 is potentially a glucose-lowering medication that can lower blood glucose, inhibit fat accumulation and ameliorate liver fibrosis.

SP8.12 - Evaluating the pig tissue model for acute proctology simulation at General Surgery ST3 Bootcamp

Abstract Evaluating the pig tissue model for acute proctology simulation at General Surgery ST3 Bootcamp Background Acute proctology is an important skill for any general surgical trainee; however it is a skill which often is overlooked when it comes to training. Most, if not all teaching happens during patient procedures. Simulation in this area is underdeveloped and there is a lack of validated models for simulation. These models are also usually expensive and difficult to obtain. The aim of this study was to assess face validity of a low-cost model for teaching acute proctology during the ST3 general surgical bootcamp. Methods We created a model using a pig rectum and perineum. This provided two uses. Firstly, we were able to teach relevant clinical anatomy. Secondly, we were able to simulate EUA, rigid sigmoidoscopy, decompression of volvulus and haemorrhoidal banding. The models allowed for repeated use. Results Twenty-five trainees took part in this simulation. Overall, the trainees felt the rectal examination was realistic (76%), the sigmoidoscope moved realistically in the model (72%) as did the proctoscope (88%). In terms of ability, trainees felt the simulation improved their confidence in EUA (68%), rectal examination (72%) and banding (80%). A total of 80% of trainees thought the model was overall “Good or Excellent”. Conclusions Our study demonstrates that this low-cost, high-fidelity simulation model has a high face validity and may be explored for the use for training new general surgical registrars.

Effect of Silica Nanoparticle Treatment on Adhesion between Tissue-like Substrates and In Vivo Skin Wound Sealing.

Silica nanoparticles are innovative solutions of surgical glue that can readily adhere to various tissue-like substrates without the need for time-consuming chemical reactions or ultraviolet irradiation. Herein, 10 nm-sized silica nanoparticle (SiNP10) treatment exhibited maximum adhesion strength in the porcine heart tissue model, which was approximately 7.15 times higher than that of the control group of non-treatment. We assessed the effects of silica nanoparticle treatment on in vivo skin wounds by scoring tissue adhesion and inflammation using histological images. Compared to the commercial cyanoacrylate skin adhesive (Dermabond), suppression of inflammatory cytokine levels in the incision wound skin was observed. We further quantified the expression of angiogenic growth factors and connective tissue formation-related proteins. On day 5 after wound closing treatment, the expression levels of PDGF-BB growth factor were significantly higher in SiNP10 treatment (0.64 ± 0.03) compared to Dermabond (0.07 ± 0.05). This stimulated angiogenesis and connective tissue formation in the skin of the incision wound may be associated with the promoting effects of SiNP10 treatment on wound closure and tissue adhesion.

REVOLUTIONIZING HEALTHCARE: THE ROLE OF NANOMATERIALS IN TISSUE ENGINEERING AND MEDICAL THERAPIES

Nanomaterials are revolutionizing tissue engineering and medical practices through their unique attributes, such as high surface area, diverse physical and chemical properties, and their ability to engage with biological systems at the molecular level. In tissue engineering, these materials significantly improve scaffold designs, which supports enhanced cell attachment, proliferation, and differentiation necessary for effective tissue regeneration. They are also crucial for the advancement of controlled drug delivery systems, which are integral to developing functional tissue models. In medical applications, nanomaterials are increasingly being used for diagnostic procedures, targeted drug delivery, and various treatments, leading to improved therapeutic outcomes and reduced side effects. Their use in imaging and diagnostics facilitates early disease detection and supports tailored treatment strategies, transforming patient care. Ongoing research into nanomaterials is expected to tackle complex healthcare issues, resulting in the development of safer and more innovative medical therapies.

Intelligent ultrasonic aspirator: Advancing tissue differentiation through hierarchical classification during hand-held resection

Modern neurosurgery strives to maximize tumor removal while preserving healthy tissue integrity. Accurate intraoperative differentiation between tumor and healthy tissue is crucial yet challenging. Often neurosurgeons rely on their experience and haptic feedback during palpation to distinguish between tumor and healthy tissue. A commonly used hand-held tool for tissue removal during neurosurgery is the ultrasonic aspirator, which changes its electrical properties as it interacts with tissue. The goal is to equip the ultrasonic aspirator with the ability to differentiate between different types of tissue while at the same time not interfering with the surgical workflow and providing comprehensible outcomes. To this end, a hierarchical classification approach is employed as a proof of concept, enabling precise identification of tissue stiffness during resection.The hierarchical approach is compared with the standard flat classification, commonly used in machine learning. Within the hierarchical approach, two strategies are employed: mandatory leaf-node predictions (MLNP) and non-mandatory leaf-node predictions (NMLNP). The NMLNP allows prediction to revert to a parent node when certainty is low. Data are acquired on three artificial tissue models – differing in stiffness – with an ultrasonic aspirator in a hand-held manner. The dataset comprises 1,821 data points for training and 186 for testing after balancing.The results indicate a slight performance advantage for the hierarchical classification MLNP approach over the flat classification approach in the absence of confidence thresholds, with weighted F2-scores of 0.781 and 0.762, respectively. However, the application of confidence thresholds results in both approaches exhibiting comparable performance, with the hierarchical NMLNP approach achieving a weighted F1-score of 0.920, thereby demonstrating superior overall performance. The effects of enforcing these thresholds and excluding data with low certainty are thoroughly investigated. This work emphasizes the feasibility of tissue differentiation using a hand-held ultrasound aspirator while resecting tissue. Moreover, it highlights the capability of hierarchical classification in advancing tissue differentiation accuracy during neurosurgical procedures, which could ultimately aid surgeons and enhance the safety of intraoperative workflows.

Bioactive Microgels with Tunable Microenvironment as a 3D Platform to Guide the Complex Physiology of Hepatocellular Carcinoma Spheroids.

Miniaturized three-dimensional tissue models, such as spheroids, have become a highly useful and efficient platform to investigate tumor physiology and explore the effect of chemotherapeutic efficacy over traditional two-dimensional monolayer culture, since they can provide more in-depth analysis, especially in regards to intercellular interactions and diffusion. The development of most tumor spheroids relies on the high proliferative capacity and self-aggregation behavior of tumor cells. However, it disregards the effect of microenvironmental factors mediated by extracellular matrix, which are indispensable components of tissue structure. In this study, hepatocellular carcinoma (HCC) cells are encapsulated in bioactive microgels consisting of gelatin and hyaluronic acid designed to emulate tumor microenvironment in order to induce hepatic tumor spheroid formation. Two different subtypes of HCC's, HepG2 and Hep3B cell lines, are explored. The physicomechanical and biochemical properties of the microgels, controlled by changing the crosslinking density and polymer composition, are clearly shown to have substantial influence over the formation and spheroid formation. Moreover, the spheroids made from different cells and microgel properties display highly variable chemoresistance effects, further highlighting the importance of microenvironmental factors guiding tumor spheroid physiology.

Efficacy of modified anterior maxillary segmental distraction osteogenesis based on 3D visualisation for the treatment of maxillary hypoplasia among adolescents with cleft lip and palate

BackgroundThis study evaluates a three-dimensional (3D) visualisation design combined with customized surgical guides to assist anterior maxillary segmental distraction osteogenesis (AMSDO) in correcting maxillary hypoplasia in adolescents with cleft lip and palate (CLP), focusing on treatment outcomes, satisfaction and the validity of 3D planning.MethodsThis retrospective cohort study was conducted at a single hospital in China. Between January 2020 and December 2023, 12 adolescents with CLP with maxillary hypoplasia were included. An advanced 3D simulation was used to convey the treatment strategy to the patients and their families. A customized surgical guide and distraction osteogenesis device were designed. Cephalometric analysis evaluated AMSDO changes and long-term stability. Patient satisfaction was assessed. The Chinese version of the Child Oral Health Impact Profile was used to evaluate the children’s oral health-related quality of life before and after treatment. The postoperative outcomes were compared with the planned outcomes by superimposing the actual postoperative data onto the simulated soft tissue models and calculating the linear and angular differences between them.ResultsOne patient experienced postoperative gingivitis, yielding an 8.33% complication rate. Most patients (83.33%) were highly satisfied with the target position, with the rest content. Cephalometric analysis showed significant improvements in various indices post-traction. Quality-of-life scores significantly improved post-treatment. The discrepancies in facial soft tissue between the simulated and actual results were within clinically satisfactory ranges.ConclusionsDigitally designed surgical guides effectively treat maxillary hypoplasia in adolescents with CLP, ensuring stability, reducing complications, reducing dependency on operator experience, and enhancing satisfaction and health outcomes. Although the simulated results were clinically acceptable, it is important to inform patients of potential variations in the predicted soft tissue.

Characterization of pediatric porcine pulmonary valves as a model for tissue engineered heart valves

Heart valve tissue engineering holds the potential to transform the surgical management of congenital heart defects affecting the pediatric pulmonary valve (PV) by offering a viable valve replacement. While aiming to recapitulate the native valve, the minimum requirement for tissue engineered heart valves (TEHVs) has historically been adequate mechanical function at implantation. However, long-term in situ functionality of TEHVs remains elusive, suggesting that a closer approximation of the native valve is required. The realization of biomimetic engineered pediatric PV is impeded by insufficient characterization of healthy pediatric tissue. In this study, we comprehensively characterized the planar biaxial tensile behaviour, extracellular matrix (ECM) composition and organization, and valvular interstitial cell (VIC) phenotypes of PVs from piglets to provide benchmarks for TEHVs. The piglet PV possessed an anisotropic and non-linear tension-strain profile from which material constants for a predictive constitutive model were derived. The ECM of the piglet PV possessed a trilayer organization populated by collagen, glycosaminoglycans, and elastin. Biochemical quantification of ECM content normalized to wet weight and DNA content of PV tissue revealed homogeneous distribution across sampled regions of the leaflet. Finally, VICs in the piglet PV were primarily quiescent vimentin-expressing fibroblasts, with a small proportion of activated α-smooth muscle actin-expressing myofibroblasts. Overall, piglet PV properties were consistent with those reported anecdotally for pediatric human PVs and distinct from those of adult porcine and human PVs, supporting the utility of the properties determined here to inform the design of tissue engineered pediatric PVs. Statement of significanceHeart valve tissue engineering has the potential to transform treatment for children born with defective pulmonary valves by providing living replacement tissue that can grow with the child. The design of tissue engineered heart valves is best informed by native valve properties, but native pediatric pulmonary valves have not been fully described to date. Here, we provide comprehensive characterization of the planar biaxial tensile behaviour, extracellular matrix composition and organization, and valvular interstitial cell phenotypes of pulmonary valves from piglets as a model for the native human pediatric valve. Together, these findings provide standards that inform engineered heart valve design towards generation of biomimetic pediatric pulmonary valves.

Multi-modal, Label-free, Optical Mapping of Cellular Metabolic Function and Oxidative Stress in 3D Engineered Brain Tissue Models.

Brain metabolism is essential for the function of organisms. While established imaging methods provide valuable insights into brain metabolic function, they lack the resolution to capture important metabolic interactions and heterogeneity at the cellular level. Label-free, two-photon excited fluorescence imaging addresses this issue by enabling dynamic metabolic assessments at the single-cell level without manipulations. In this study, we demonstrate the impact of spectral imaging on the development of rigorous intensity and lifetime label-free imaging protocols to assess dynamically over time metabolic function in 3D engineered brain tissue models comprising human induced neural stem cells, astrocytes, and microglia. Specifically, we rely on multi-wavelength spectral imaging to identify the excitation/emission profiles of key cellular fluorophores within human brain cells, including NAD(P)H, LipDH, FAD, and lipofuscin. These enable development of methods to mitigate lipofuscin's overlap with NAD(P)H and flavin autofluorescence to extract reliable optical metabolic function metrics from images acquired at two excitation wavelengths over two emission bands. We present fluorescence intensity and lifetime metrics reporting on redox state, mitochondrial fragmentation, and NAD(P)H binding status in neuronal monoculture and triculture systems, to highlight the functional impact of metabolic interactions between different cell types. Our findings reveal significant metabolic differences between neurons and glial cells, shedding light on metabolic pathway utilization, including the glutathione pathway, OXPHOS, glycolysis, and fatty acid oxidation. Collectively, our studies establish a label-free, non-destructive approach to assess the metabolic function and interactions among different brain cell types relying on endogenous fluorescence and illustrate the complementary nature of information that is gained by combining intensity and lifetime-based images. Such methods can improve understanding of physiological brain function and dysfunction that occurs at the onset of cancers, traumatic injuries and neurodegenerative diseases.

Dynamic monitoring of a 3D-printed airway tissue model using an organic electrochemical transistor

Assessing the transepithelial resistance to ion flow in the presence of an electric field enables the evaluation of the integrity of the epithelial cell layer. In this study, we introduce an organic electrochemical transistor (OECT) interfaced with a 3D living tissue, designed to monitor the electrical resistance of cellular barriers in real-time. We have developed a non-invasive, tissue-sensing platform by integrating an inkjet-printed large-area OECT with a 3D-bioprinted multilayered airway tissue. This unique configuration enables the evaluation of epithelial barrier integrity through the dynamic response capabilities of the OECT. Our system effectively tracks the formation and integrity of 3D-printed airway tissues in both liquid-liquid and air-liquid interface culture environments. Furthermore, we successfully quantified the degradation of barrier function due to influenza A (H1N1) viral infection and the dose-dependent efficacy of oseltamivir (Tamiflu®) in mitigating this degradation. The tissue-electronic platform offers a non-invasive and label-free method for real-time monitoring of 3D artificial tissue barriers, without disturbing the cellular biology. It holds the potential for further applications in monitoring the structures and functions of 3D tissues and organs, significantly contributing to the advancement of personalized medicine.

Synergistic effects of biological stimuli and flexion induce microcavities promote hypertrophy and inhibit chondrogenesis during in vitro culture of human mesenchymal stem cell aggregates.

Interzone/cavitation are key steps in early stage joint formation that have not been successfully developed in vitro. Further, current models of endochondral ossification, an important step in early bone formation, lack key morphology morphological structures such as microcavities found during development in vivo. This is possibly due to the lack of appropriate strategies for incorporating chemical and mechanical stimuli that are thought to be involved in joint development. We designed a bioreactor system and investigated the synergic effect of chemical stimuli (chondrogenesis-inducing [CIM] and hypertrophy-inducing medium [HIM]) and mechanical stimuli (flexion) on the growth of human mesenchymal stem cells (hMSCs) based linear aggregates under different conditions over 4 weeks of perfusion culture. Computational studies were used to evaluate tissue stress qualitatively. After harvesting, both Safranin-O and hematoxylin & eosin (H&E) staining histology demonstrated microcavity structures and void structures in the region of higher stresses for tissue aggregates cultured only in HIM under flexion. In comparison to either HIM treatment or flexion only, increased glycosaminoglycan (GAG) content in the extracellular matrix (ECM) at this region indicates the morphological change resembles the early stage of joint cavitation; while decreased type II collagen (Col II), and increased type X collagen (Col X) and vascular endothelial growth factor (VEGF) with a clear boundary in the staining section indicates it resembles the early stage of ossification. Further, cell alignment analysis indicated that cells were mostly oriented toward the direction of flexion in high-stress region only in HIM under flexion, resembling cell morphology in both joint cavitation and hypertrophic cartilage in growth plate. Collectively, our results suggest that flexion and HIM inhibit chondrogenesis and promote hypertrophy and development of microcavities that resemble the early stage of joint cavitation and endochondral ossification. We believe the tissue model described in this work can be used to develop in vitro models of joint tissue for applications such as pathophysiology and drug discovery.

High-Speed Thermal Imaging Can Resolve Short RF Pulse Effects in Tissue Models

High-Speed Thermal Imaging Can Resolve Short RF Pulse Effects in Tissue Models

Development of Sensitive In Vitro Protocols for the Biocompatibility Testing of Medical Devices and Pharmaceuticals Intended for Contact with the Eyes: Acute Irritation and Phototoxicity Assessment.

This study introduces a novel in vitro methodology that employs the 3-D reconstructed tissue model, EpiOcular, to assess the irritation and phototoxicity potential of medical devices and drugs in contact with the eye. Our study evaluated diverse test materials, including medical devices, ophthalmological solutions and an experimental drug (cemtirestat), for their potential to cause eye irritation and phototoxicity. The protocols used in this study with the EpiOcular tissue model were akin to those used in the ultra-mildness testing of cosmetic formulations, which is challenging to predict with standard in vivo rabbit tests. To design these protocols, we leveraged experience gained from the validation project on the EpiDerm skin irritation test for medical devices (ISO 10993-23:2021) and the OECD TG 498 method for photo-irritation testing. The predictions were based on the tissue viability and inflammatory response, as determined by IL-1α release. By developing and evaluating these protocols for medical devices, we aimed to expand the applicability domain of the tests referred to in ISO 10993-23. This will contribute to the standardisation and cost-effective safety evaluation of ophthalmic products, while reducing reliance on animal testing in this field. The findings obtained from the EpiOcular model in the photo-irritation test could support its implementation in the testing strategies outlined in OECD TG 498.