Tissue Model Research Articles

T Cells Spatially Regulate B Cell Receptor Signaling in Lymphomas through H3K9me3 Modifications.

Activated B cell-like diffuse large B-cell lymphoma (ABC-DLBCL) is a subtype associated with poor survival outcomes. Despite identifying therapeutic targets through molecular characterization, targeted therapies have limited success. New strategies using immune-competent tissue models are needed to understand how DLBCL cells evade treatment. Here, synthetic hydrogel-based lymphoma organoids are used to demonstrate how signals in the lymphoid tumor microenvironment (Ly-TME) can alter B cell receptor (BCR) signaling and specific histone modifications, tri-methylation of histone 3 at lysine 9 (H3K9me3), dampening the effects of BCR pathway inhibition. Using imaging modalities, T cells increase DNA methyltransferase 3A expression and cytoskeleton formation in proximal ABC-DLBCL cells, regulated by H3K9me3. Expansion microscopy on lymphoma organoids reveals T cells increase the size and quantity of segregated H3K9me3 clusters in ABC-DLBCL cells. Findings suggest the re-organization of higher-order chromatin structures that may contribute to evasion or resistance to therapy via the emergence of novel transcriptional states. Treating ABC-DLBCL cells with a G9α histone methyltransferase inhibitor reverses T cell-mediated modulation of H3K9me3 and overcomes T cell-mediated attenuation of treatment response to BCR pathway inhibition. This study emphasizes the Ly-TME's role in altering DLBCL fate and suggests targeting aberrant signaling and microenvironmental cross-talk that can benefit high-risk patients.

Exploring the bias: how skin color influences oxygen saturation readings via Monte Carlo simulations.

Our goal is to understand the root cause of reported oxygen saturation ( ) overestimation in heavily pigmented skin types to devise solutions toward enabling equity in pulse oximeter designs. We aim to gain theoretical insights into the effect of skin tone on curves using a three-dimensional, four-layer tissue model representing a finger. A finger tissue model, comprising the epidermis, dermis, two arteries, and a bone, was developed using a Monte Carlo-based approach in the MCmatlab software. Two skin tones-light and dark-were simulated by adjusting the absorption and scattering properties within the epidermal layer. Following this, curves were generated in various tissue configurations, including transmission and reflection modes using red and infrared wavelengths. In addition, the influence of source-detector (SD) separation distances on both light and dark skin tissue models was studied. In transmission mode, curves did not deviate with changes in skin tones because both pulsatile and non-pulsatile terms experienced equal attenuation at red and infrared wavelengths. However, in reflection mode, measurable variations in curves were evident. This was due to differential attenuation of the red components, which resulted in a lower perfusion index at the red wavelength in darker skin. As the SD separation increased, the effect of skin tone on curves in reflection mode became less pronounced, with the largest SD separation exhibiting effects similar to those observed in transmission mode. Monte Carlo simulations have demonstrated that different light pathlengths within the tissue contribute to the overestimation of in people with darker skin in reflection mode pulse oximetry. Increasing the SD separation may mitigate the effect of skin tone on readings. These trends were not observed in transmission mode; however, further planned research using more complex models of the tissue is essential.

Vaginal irritation testing-prospects of human organotypic vaginal tissue culture models.

Personal lubricants intended for local or systemic delivery via the vaginal route can induce vaginal irritation, damage the vaginal epithelial barrier which can enhance microbial entry, induce inflammation, and alter the microbiome of the vaginal ecosystem. Therefore, manufacturers of personal lubricants and medical devices are required to show biocompatibility and safety assessment data to support regulatory decision-making within a specified context of use. Furthermore, due to ethical concerns and the introduction of the 7th amendment of the European Council Directive which bans animal testing for cosmetic ingredients and products coupled with the Food and Drug Administration modernization Act 2.0 guidelines, there is a wave of drive to develop alternative test methods to predict human responses to chemical or formulation exposure. In this framework, there is a potential to use three-dimensional organotypic human vaginal-ectocervical tissue models as a screening tool to predict the vaginal irritation potential of personal lubricants and medicaments. To be physiologically relevant, the in vitro tissue models need to be reconstructed using primary epithelial cells of the specific organ or tissue and produce organ-like structure and functionality that recapitulate the in vivo-like responses. Through the years, progress has been made and vaginal tissue models are manufactured under controlled conditions with a specified performance criterion, which leads to a high level of reproducibility and reliability. The utility of vaginal tissue models has been accelerated in the last 20years with an expanded portfolio of applications ranging from toxicity, inflammation, infection to drug safety, and efficacy studies. This article provides an overview of the state of the art of diversified applications of reconstructed vaginal tissue models and highlights their utility as a tool to predict vaginal irritation potential of feminine care products.

Advancing diagnostics and disease modeling: current concepts in biofabrication of soft microfluidic systems

Soft microfluidic systems play a pivotal role in personalized medicine, particularly in in vitro diagnostics tools and disease modeling. These systems offer unprecedented precision and versatility, enabling the creation of intricate three-dimensional (3D) tissue models that can closely emulate both physiological and pathophysiological conditions. By leveraging innovative biomaterials and bioinks, soft microfluidic systems can circumvent the current limitations involving the use of polydimethylsiloxane (PDMS), thus facilitating the development of customizable systems capable of sustaining the functions of encapsulated cells and mimicking complex biological microenvironments. The integration of lab-on-a-chip technologies with soft nanodevices further enhances disease models, paving the way for tailored therapeutic strategies. The current research concepts underscore the transformative potential of soft microfluidic systems, exemplified by recent breakthroughs in soft lithography and 3D (bio)printing. Novel applications, such as multi-layered tissues-on-chips and skin-on-a-chip devices, demonstrate significant advancements in disease modeling and personalized medicine. However, further exploration is warranted to address challenges in replicating intricate tissue structures while ensuring scalability and reproducibility. This exploration promises to drive innovation in biomedical research and healthcare, thus offering new insights and solutions to complex medical challenges and unmet needs.

Immunocompetent Alveolus-on-Chip Model for Studying Alveolar Mucosal Immune Responses.

We introduce an advanced immunocompetent lung-on-chip model designed to replicate the human alveolar structure and function. This innovative model employs a microfluidic-perfused biochip that supports an air-liquid interface mimicking the environment in the human alveoli. Tissue engineering is used to integrate key cellular components, including endothelial cells, macrophages, and epithelial cells, to create a representative tissue model of the alveolus. The model facilitates in-depth examinations of the mucosal immune responses to various pathogens, including viruses, bacteria, and fungi, thereby advancing our understanding of lung immunity. The primary goal of this protocol is to provide details for establishing this alveolus-on-chip model as a robust in vitro platform for infection studies, enabling researchers to closely observe and analyze the complex interactions between pathogens and the host's immune system within the pulmonary environment. This is achieved through the application of microfluidic-based techniques to simulate key physiological conditions of the human alveoli, including blood flow and biomechanical stimulation of endothelial cells, alongside maintaining an air-liquid interface crucial for the realistic exposure of epithelial cells to air. The model system is compatible with a range of standardized assays, such as immunofluorescence staining, cytokine profiling, and colony-forming unit (CFU)/plaque analysis, allowing for comprehensive insights into immune dynamics during infection. The Alveolus-on-chip is composed of essential cell types, including human distal lung epithelial cells (H441) and human umbilical vein endothelial cells (HUVECs) separated by porous polyethylene terephthalate (PET) membranes, with primary monocyte-derived macrophages strategically positioned between the epithelial and endothelial layers. The tissue model enhances the ability to dissect and analyze the nuanced factors involved in pulmonary immune responses in vitro. As a valuable tool, it should contribute to the advancement of lung research, providing a more accurate and dynamic in vitro model for studying the pathogenesis of respiratory infections and testing potential therapeutic interventions.

Particle Detection in Free-Falling Nanoliter Droplets.

Sorting and dispensing distinct numbers of cellular aggregates enables the creation of three-dimensional (3D) in vitro models that replicate in vivo tissues, such as tumor tissue, with realistic metabolic properties. One method for creating these models involves utilizing Drop-on-Demand (DoD) dispensing of individual Multicellular Spheroids (MCSs) according to material jetting processes. In the DoD approach, a droplet dispenser ejects droplets containing these MCSs. For the reliable printing of tissue models, the exact number of dispensed MCSs must be determined. Current systems are designed to detect MCSs in the nozzle region prior to the dispensing process. However, due to surface effects, in some cases the spheroids that are detected adhere to the nozzle and are not dispensed with the droplet as expected. In contrast, detection that is carried out only after the droplet has been ejected is not affected by this issue. This work presents a system that can detect micrometer-sized synthetic or biological particles within free-falling droplets with a volume of about 30 nanoliters. Different illumination modalities and detection algorithms were tested. For a glare point projection-based approach, detection accuracies of an average of 95% were achieved for polymer particles and MCF-7 spheroids with diameters above 75 μm. For smaller particles the detection accuracy was still in the range of 70%. An approach with diffuse white light illumination demonstrated an improvement for the detection of small opaque particles. Accuracies up to 96% were achieved using this concept. This makes the two demonstrated methods suitable for improving the accuracy and quality control of particle detection in droplets for Drop-on-Demand techniques and for bioprinting.

Modelling of a RYR2-associated left ventricular non-compaction and sudden cardiac death overlap syndrome in hiPSC-EHTs

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): ESC training grant; DZHK Background Genetic cardiomyopathies contribute to a significant proportion of morbidity and mortality among young people. Over the past decades, a wide range of underlying molecular mechanisms has been discovered. The current project arose from the clinical observation of a left ventricular non-compaction and sudden cardiac death overlap syndrome in a large family in northern Italy. Whole exome sequencing performed on family members of different generations revealed a single nucleotide polymorphism in the RYR2 gene (RYR2 c.5654G>A homozygous – RyR2 p.G1885E - exon37:missense) with an autosomal-recessive inheritance pattern. The mutation is localized in the DR3 region of the RyR2 protein, adjacent to the FKBP12.6 binding site, a protein that is known to stabilize RyR2, preventing aberrant activation of the channel. Methods and Results Patient-derived, human induced pluripotent stem cells (hiPSCs), carrying the variant homo- and heterozygously, were generated by reprogramming patient-derived peripheral blood mononuclear cells using a non-integrative Sendai virus protocol. The lines were fully characterized and expressed high levels of pluripotency markers at the RNA and protein level. Patient lines were all able to efficiently differentiate into cardiomyocytes. Subsequently, 3D strip-format, force-generating Engineered Heart Tissues (EHTs) were cast and structurally and functionally characterized. Contraction analysis over 60 days showed differences in force development of the patient-derived lines. In particular, the EHTs carrying the homozygous pathogenic variant showed progressive reduction in both beating rate and force over time, increased spontaneous arrhythmogenicity and reduced inotropic response after exposure to isoprenaline, compared to the heterozygous line and unrelated healthy controls. Starting from day 30 after casting, patient-derived EHTs developed morphological changes with areas of marked cellular overgrowth and reduced troponin expression. Transcriptomic and proteomic analysis at 30 and 60 days after EHTs generation, as well as in 2D cardiomyocytes, complement the phenotyping and provide molecular insights into the underlying pathophysiology. To directly evaluate pathogenicity of the genetic variant identified, we generated isogenic homo- and heterozygously corrected hiPSCs, using CRISPR/ Cas9 technology, starting from the homozygous patient-line. The characterization of these corrected clones is currently ongoing. The genetically corrected 3D tissue models will be critical not only in defining the role of the novel RyR2 variant in the familial phenotype described, but also to unveil mechanistic roles of ryanodine receptor in cardiac disease. Conclusion We describe a novel recessive RYR2 variant associated with a left ventricular non- compaction and sudden cardiac death overlap syndrome. 3D cardiac tissue modelling based on patient-derived hiPSCs uncovered a proarrhythmic and structural phenotype of the mutated lines.

Modeling doxorubicin-induced cardiotoxicity with human myocardial tissue slices: insights on genetic susceptibility and cardioprotection

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): PSIDER and HARVEY (ZonMW) Introduction Cardiotoxicity poses a significant challenge in drug development, often leading to the withdrawal of approved drugs. To address this issue, there is a critical need for improved models to predict cardiac adverse effects. In line with the imperative for animal-free preclinical testing, this study aimed to develop a novel model using human myocardial tissue slices to get a deeper understanding and potential prevention of doxorubicin-Induced cardiotoxicity. Methods Human myocardial tissue slices of 300 μm thick were derived from end-stage heart failure patients explanted hearts, including those harbouring likely or pathogenic cardiovascular mutations (class 4 and 5) (N=3) and non-genetic cardiovascular etiology (N=3). Myocardial tissue slices were cultured under physiological mechanical and electrical conditions. Slices were exposed to doxorubicin (1 µM) over a 10-day culture period, and their responses were compared to DMSO-vehicle controls, by recording contractile force and Ca2+ transients. Additionally, a subset of slices was treated with a cardioprotective agent, dexrazoxane (100 µM), 1 day prior to and during doxorubicin exposure to assess potential cardioprotection. Results Doxorubicin-exposed slices with a pathogenic mutation exhibited reduced mechanical force, increased threshold potential, and impaired ability to follow pacing at higher frequencies. Additionally, aberrations in calcium handling, manifested as arrhythmia-like disconnection of muscle fibers, were notably observed in these slices. Instead, slices from patients without pathogenic mutations showed minimal cardiotoxicity when exposed to doxorubicin. Encouragingly, pretreatment with dexrazoxane demonstrated a protective effect against doxorubicin-induced cardiotoxicity in the genetically susceptible group. Our functional findings corroborated our structural data, highlighting the breakdown of sarcomere structure and cell-cell connections and increased DNA damage, particularly in patients harbouring a genetic cardiovascular mutation. Conclusion In summary, human myocardial tissue slices faithfully recapitulate doxorubicin-induced cardiotoxicity. Importantly, our data suggests the utility of employing genetic testing for known cardiovascular mutations as a valuable step before initiating doxorubicin treatment. Additionally, we demonstrate for the first time the in vitro cardioprotective potential of dexrazoxane in a human tissue model. These findings pave the way for using ex vivo myocardial tissue slices to evaluate cardiotoxicity and their value as a powerful tool for evaluating the efficacy and delivery approaches.

What is the weakest point of a secured aortic cannula in central extracorporeal membrane oxygenation?

The purpose of this study was to compare techniques for securing the aortic extracorporeal membrane oxygenation (ECMO) cannula, using in vitro models. Two models were studied: a tissue model using porcine aortas and a stand model replacing the aorta with a metal stand to study the system independent of the tissue. Interventions in each model were divided into three experimental groups: Group 1 (3-0 Prolene® + 20-French Medtronic Arterial Cannula EOPA™), Group 2 (4-0 Prolene® + 16-French Medtronic Arterial Cannula DLP Pediatric), and Group 3 (5-0 Prolene® + 8-French Medtronic Arterial Cannula DLP Pediatric). In separate experiments, both gradual and rapid forces were applied to the cannulas, starting with 9.8 Newtons and increasing exponentially if the cannula remained secured. Additionally, the method of securing the tourniquet and the number of ties securing the tourniquet to the cannula were evaluated. In the tissue model, even with a minimum force of 9.8 Newtons, the suture pulled through the aortic tissue, leaving sutures and ties intact. In the stand model, two purse-string sutures secured by two ligaclips held the cannula reliably and withstood higher total force. Dislodgement was prevented at forces close to 60 Newtons with only two hemostatic clips included in cannulation. The weakest part of the aortic ECMO cannulation system using in vitro experiments was the tissue. Assuming that these experiments translate in vivo, it is therefore critical to prevent any pull on the cannulas by securing ECMO cannulas and ECMO tubing to both the patient and the patient's bed. Sutures with a larger diameter withstand more force. Two medium hemostatic clips can secure Prolene® sutures within snares as safely as a mosquito hemostat. Two polypropylene purse-string sutures secured by two hemostatic clips were most reliable at greater forces. The rationale for publishing our experiments in this manuscript is to (1) communicate our quantification of possible contributing factors to this rare and likely catastrophic complication of unintended decannulation, (2) increase awareness about this potential complication, and (3) increase vigilance to assure prevention of this dreaded complication.

Vascular Anastomoses and Dissection: A Six-Part Simulation Curriculum for Surgical Residents.

As surgical technologies grow, so too do demands on surgical trainees to master increasing numbers of skill sets. With the rise of endovascular surgery, trainees have fewer opportunities to practice open vascular techniques in the operating room. Simulation can bridge this gap. However, existing published open vascular simulation curricula are basic or based on expensive models. We iteratively developed an open vascular skills curriculum for second-year surgery residents comprising six 2-hour sessions. We refined the curriculum based on feedback from learners and faculty. The curriculum required skilled facilitators, vascular instruments, and tissue models. We evaluated the latest iteration with a survey and by assessing participants' technical skills using the Objective Structured Assessment of Technical Skills (OSATS) form. Over the past 10 years, 101 residents have participated in the curriculum. Nine of 13 residents who participated in the latest curricular iteration completed the survey. All respondents rated the sessions as excellent and strongly agreed that they had improved their abilities to perform anastomoses with tissue and prosthetic. Facilitators completed 18 OSATS forms for residents in the fifth and sixth sessions of the latest iteration. Residents scored well overall, with a median 26.5 (interquartile range: 24-29) out of a possible score of 35, with highest scores on knowledge of instruments. This simulation-based curriculum facilitates open vascular surgical skill acquisition among surgery residents. The curriculum allows residents to acquire critical vascular skills that are challenging to learn in an increasingly demanding operative setting.

Cytotoxicity and Bioimmunological Activity of Poly(2-Isopropenyl-2-oxazoline) Conjugates with Ibuprofen Using 3D Reconstructed Tissue Models.

Poly(2-isopropenyl-2-oxazoline) (PIPOx) represents a universal polymer platform with pendant 2-oxazoline groups, allowing the preparation of biomaterials for various biomedical applications. However, there is a lack of information on PIPOx concerning the effect of molar mass (Mn) on cytotoxicity and bioimmunological properties. Here, aqueous copper(0)-mediated reversible-deactivation radical polymerization (Cu0-RDPR) was used for the preparation of PIPOx with defined Mn and low dispersity. PIPOx of different Mn are used for the synthesis of conjugates with ibuprofen (5 mol %), the nonsteroidal anti-inflammatory drug. The release of ibuprofen at 37 °C and different pH values is monitored using high-performance liquid chromatography, where the rate of drug release increases with increasing pH and lower Mn. In vitro cytotoxicity and bioimmunological properties of PIPOx and drug conjugates are studied using 3D reconstructed tissue models of the human epidermis and intestinal epithelium. We demonstrate low cytotoxicity of PIPOx and conjugates with different Mn values on both 3D tissue models.

Natural elicitors enhanced suberin polyphenolic accumulation in wounded potato tuber tissues.

Unintended wounding or bruising during harvest or postharvest handling leads to significant tuber loss and imposes economic burden to potato industry. Therefore, finding effective strategies to mitigate wound-related tuber losses is very important from industry perspectives. Formation of protective barrier through accumulation of suberin polyphenolics (SPP) is a natural and initial response of potato tuber tissues to wounding. In this study, efficacy of two natural elicitors, such as chitosan oligosaccharide (COS 0.125 g L-1) and cranberry pomace residue (Nutri-Cran 0.125 g L-1) was investigated using a mechanically wounded tuber tissue model and by histological determination of SPP formation in five agronomically relevant and red-skin potato cultivars (Chieftain, Dakota Rose, Dakota Ruby, Red LaSoda, Red Norland). Furthermore, the potential role of stress protective metabolic regulation involving phenolic metabolites, proline, and antioxidant enzymes in tuber WH processes were also investigated during 0-9 days after wounding. Exogenous treatments of both COS and Nutri-Cran resulted into enhanced SPP formation in wounded surface, but the impact was more rapid with Nutri-Cran treatment in select cultivars. Greater contents of total soluble phenolic, ferulic acid, chlorogenic acid, total antioxidant activity, and superoxide dismutase activity were also observed in elicitor treated tuber tissues at different time points after wounding. Nutri-Cran treatment also reduced the activity of succinate dehydrogenase in Red Norland and Dakota Ruby at 3 d, indicating a suppression in respiration rate. Collectively, these results suggest that Nutri-Cran can be potentially utilized as an effective WH treatment to potato tubers for minimizing wound-related losses.

Toward a disruptive, minimally invasive small finger joint implant concept: Cellular and molecular interactions with materials in vivo

Osteoarthritis (OA) poses significant therapeutic challenges, particularly OA that affects the hand. Currently available treatment strategies are often limited in terms of their efficacy in managing pain, regulating invasiveness, and restoring joint function. The APRICOTⓇ implant system developed by Aurora Medical Ltd (Chichester, UK) introduces a minimally invasive, bone-conserving approach for treating hand OA (https://apricot-project.eu/). By utilizing polycarbonate urethane (PCU), this implant incorporates a caterpillar track-inspired design to promote the restoration of natural movement to the joint. Surface modifications of PCU have been proposed for the biological fixation of the implant. This study investigated the biocompatibility of PCU alone or in combination with two surface modifications, namely dopamine-carboxymethylcellulose (dCMC) and calcium-phosphate (CaP) coatings. In a rat soft tissue model, native and CaP-coated PCU foils did not increase cellular migration or cytotoxicity at the implant–soft tissue interface after 3 d, showing gene expression of proinflammatory cytokines similar to that in non-implanted sham sites. However, dCMC induced an amplified initial inflammatory response that was characterized by increased chemotaxis and cytotoxicity, as well as pronounced gene activation of proinflammatory macrophages and neoangiogenesis. By 21 d, inflammation subsided in all the groups, allowing for implant encapsulation. In a rat bone model, 6 d and 28 d after release of the periosteum, all implant types were adapted to the bone surface with a surrounding fibrous capsule and no protracted inflammatory response was observed. These findings demonstrated the biocompatibility of native and CaP-coated PCU foils as components of APRICOTⓇ implants. Statement of significanceHand osteoarthritis treatments require materials that minimize irritation of the delicate finger joints. Differing from existing treatments, the APRICOTⓇ implant leverages polycarbonate urethane (PCU) for minimally invasive joint replacement. This interdisciplinary, preclinical study investigated the biocompatibility of thin polycarbonate urethane (PCU) foils and their surface modifications with calcium-phosphate (CaP) or dopamine-carboxymethylcellulose (dCMC). Cellular and morphological analyses revealed that both native and Ca-P coated PCU elicit transient inflammation, similar to sham sites, and a thin fibrous encapsulation in soft tissues and on bone surfaces. However, dCMC surface modification amplified initial chemotaxis and cytotoxicity, with pronounced activation of proinflammatory and neoangiogenesis genes. Therefore, native and CaP-coated PCU possess sought-for biocompatible properties, crucial for patient safety and performance of APRICOTⓇ implant.

Simultaneous assessment of blood flow and myelin content in the brain white matter with dynamic [11C]PiB PET: a test-retest study in healthy controls.

Exploring the relationship between oxygen supply and myelin damage would benefit from a simultaneous quantification of myelin and cerebral blood flow (CBF) in the brain's white matter (WM). To validate an analytical method for quantifying both CBF and myelin content in the WM using dynamic [11C]PiB positron emission tomography (PET). A test-retest study was performed on eight healthy subjects who underwent two consecutive dynamic [11C]PiB-PET scans. Three quantitative approaches were compared: simplified reference tissue model 2 (SRTM2), LOGAN graphical model, and standardized uptake value ratio (SUVR). The sensitivity of methods to the size of the region of interest was explored by simulating lesion masks obtained from 36 subjects with multiple sclerosis. Reproducibility was assessed using the relative difference and interclass correlation coefficient. Repeated measures correlations were used to test for cross-correlations between metrics. Among the CBF measures, the relative delivery (R1) of the simplified reference tissue model 2 (SRTM2) displayed the best reproducibility in the white matter, with a strong influence of the size of regions analyzed, the test-retest variability being below 10% for regions above 68 mm3 in the supratentorial white matter. [11C]PiB PET-derived proxies of CBF demonstrated lower perfusion of white matter compared to grey matter with an overall ratio equal to 1.71 ± 0.09 when the SRTM2-R1 was employed. Tissue binding in the white matter was well estimated by the Logan graphical model through estimation of the distribution volume ratio (LOGAN-DVR) and SRTM2 distribution volume ratio (SRTM2-DVR), with test-retest variability being below 10% for regions exceeding 106 mm3 for LOGAN-DVR and 300 mm3 for SRTM2-DVR. SRTM2-DVR provided a better contrast between white matter and grey matter. The interhemispheric variability was also dependent on the size of the region analyzed, being below 10% for regions above 103 mm3 for SRTM2-R1 and above 110 mm3 for LOGAN-DVR. Whereas the 1 to 8-minute standardized uptake value ratio (SUVR1-8) showed an intermediary reproducibility for CBF assessment, SUVR0-2 for perfusion or SUVR50-70 for tissue binding showed poor reproducibility and correlated only mildly with SRTM2-R1 and LOGAN-DVR estimations respectively. [11C]PiB PET imaging can simultaneously quantify perfusion and myelin content in WM diseases associated with focal lesions. For longitudinal studies, SRTM2-R1 and DVR should be preferred over SUVR for the assessment of regional CBF and myelin content, respectively. European Union Clinical Trials Register EUDRACT; EudraCT Number: 2008-004174-40; Date: 2009-03-06; https//www.clinicaltrialsregister.eu ; number 2008-004174-40.

In vitro throughput screening of anticancer drugs using patient-derived cell lines cultured on vascularized three-dimensional stromal tissues

The development of high-throughput anticancer drug screening methods using patient-derived cancer cell (PDC) lines that maintain their original characteristics in an in vitro three-dimensional (3D) culture system poses a significant challenge to achieving personalized cancer medicine. Because stromal tissue plays a critical role in the composition and maintenance of the cancer microenvironment, in vitro 3D-culture using reconstructed stromal tissues has attracted considerable attention. Here, a simple and unique in vitro 3D-culture method using heparin and collagen together with fibroblasts and endothelial cells to fabricate vascularized 3D-stromal tissues for in vitro culture of PDCs is reported. Whereas co-treatment with bevacizumab, a monoclonal antibody against vascular endothelial growth factor, and 5-fluorouracil significantly reduced the survival rate of 3D-cultured PDCs to 30%, separate addition of each drug did not induce comparable strong cytotoxicity, suggesting the possibility of evaluating the combined effect of anticancer drugs and angiogenesis inhibitors. Surprisingly, drug evaluation using eight PDC lines with the 3D-culture method resulted in a drug efficacy concordance rate of 75% with clinical outcomes. The model is expected to be applicable to in vitro throughput drug screening for the development of personalized cancer medicine. Statement of significanceTo replicate the cancer microenvironment, we constructed a cancer-stromal tissue model in which cancer cells are placed above and inside stromal tissue with vascular network structures derived from vascular endothelial cells in fibroblast tissue using CAViTs method. Using this method, we were able to reproduce the invasion and metastasis processes of cancer cells observed in vivo. Using patient-derived cancer cells, we assessed the possibility of evaluating the combined effect with an angiogenesis inhibitor.Further, primary cancer cells also grew on the stromal tissues with the normal medium. These data suggest that the model may be useful for new in vitro drug screening and personalized cancer medicine.

Enhancing Corneal Drug Penetration Using Penetratin for Ophthalmic Suspensions.

Eye drops, including solutions and suspensions, are essential dosage forms to treat ophthalmic diseases, with poorly water-soluble drugs typically formulated as ophthalmic suspensions. In addition to low bioavailability, suspensions exhibit limited efficacy, safety, and usability due to the presence of drug particles. Improving bioavailability can reduce the drug concentrations and the risk of problems associated with suspended drug particles. However, practical penetration enhancers capable of improving bioavailability remain elusive. Herein, we focused on penetratin (PNT), a cell-penetrating peptide (CPP) that promotes active cellular transport related to macromolecule uptake, such as micropinocytosis. According to the in vitro corneal uptake study using a reconstructed human corneal epithelial tissue model, LabCyte CORNEA-MODEL24, PNT enhanced the uptake of Fluoresbrite® YG carboxylate polystyrene microspheres without covalent binding. In an ex vivo porcine eye model, the addition of 10 µM PNT to rebamipide ophthalmic suspension markedly improved the corneal uptake of rebamipide; however, the addition of 100 µM PNT was ineffective due to potentially increased particle size by aggregation. This article provides basic information on the application of PNT as a penetration enhancer in ophthalmic suspensions, including the in vitro and ex vivo studies mentioned above, as well as the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assay and storage stability at different pH values.

SIMULATION AND EX VIVO EXPERIMENTAL STUDY OF MICROWAVE ABLATION OF ADRENAL ADENOMA BY COAXIAL SLOT ANTENNA

Object: Microwave ablation has been widely used in tumor treatment, and the antenna is a critical component of microwave ablation. We aimed to evaluate a single-slot antenna to generate a small volume ablation zone ([Formula: see text] in adrenal tissue. Methods: A three-dimensional numerical model of the adrenal tissue was built and solved by the Finite Element Method. In the simulation model, the slot antenna was placed between the adrenal gland and fat, and the microwave ablation power and time were adjusted to obtain the target ablation zone. In ex vivo experiments, the length and width of the ablation zone were measured, and the significance of ablation parameters on it was determined using a multi-factorial analysis of variance method. The temperature versus time curves at 3[Formula: see text]mm and 7[Formula: see text]mm from the antenna were plotted to validate the simulation. Results: The simulation results showed that the combination of microwave ablation parameter settings 30[Formula: see text]W/300[Formula: see text]s, 40[Formula: see text]W/180[Formula: see text]s, and 40[Formula: see text]W/300[Formula: see text]s could create ablation zones in the adrenal tissue volumes of [Formula: see text] and [Formula: see text], respectively, with minimal damage (less than [Formula: see text] to the surrounding fat tissue. Both power and heating time significantly affected the length and width of the ablation zone. The temperature rise curves obtained in the simulation and ex vivo experiments were in good agreement. Conclusion: This work shows that utilizing a slot antenna can produce a small volume ablation zone in adrenal tissue for microwave ablation of benign adrenal adenomas.

Guidelines for Manufacturing and Application of Organoids: Brain.

This study offers a comprehensive overview of brain organoids for researchers. It combines expert opinions with technical summaries on organoid definitions, characteristics, culture methods, and quality control. This approach aims to enhance the utilization of brain organoids in research. Brain organoids, as three-dimensional human cell models mimicking the nervous system, hold immense promise for studying the human brain. They offer advantages over traditional methods, replicating anatomical structures, physiological features, and complex neuronal networks. Additionally, brain organoids can model nervous system development and interactions between cell types and the microenvironment. By providing a foundation for utilizing the most human-relevant tissue models, this work empowers researchers to overcome limitations of two-dimensional cultures and conduct advanced disease modeling research.

Characterization of a Human Respiratory Mucosa Model to Study Odorant Metabolism.

Nasal xenobiotic metabolizing enzymes (XMEs) are important for the sense of smell because they influence odorant availability and quality. Since the major part of the human nasal cavity is lined by a respiratory mucosa, we hypothesized that this tissue contributed to nasal odorant metabolism through XME activity. Thus, we built human respiratory tissue models and characterized the XME profiles using single-cell RNA sequencing. We focused on the XMEs dicarbonyl and l-xylulose reductase, aldehyde dehydrogenase (ALDH) 1A1, and ALDH3A1, which play a role in food odorant metabolism. We demonstrated protein abundance and localization in the tissue models and showed the metabolic activity of the corresponding enzyme families by exposing the models to the odorants 3,4-hexandione and benzaldehyde. Using gas chromatography coupled with mass spectrometry, we observed, for example, a significantly higher formation of the corresponding metabolites 4-hydroxy-3-hexanone (39.03 ± 1.5%, p = 0.0022), benzyl alcohol (10.05 ± 0.88%, p = 0.0008), and benzoic acid (8.49 ± 0.57%, p = 0.0004) in odorant-treated tissue models compared to untreated controls (0 ± 0, 0.12 ± 0.12, and 0.18 ± 0.18%, respectively). This is the first study that reveals the XME profile of tissue-engineered human respiratory mucosa models and demonstrates their suitability to study nasal odorant metabolism.

Оценка накопления полимерных субмикронных микрокапсул в клеточном и межклеточном пространствах 3D сфероидов

Multicellular spheroids are three-dimensional in vitro models of organs and tissues. Multicellular spheroids have gained great interest in the field of biotechnology as they are reproducible and mimic real organs and tissues so can be used as test systems for new forms of drugs, allowing minimizing the use of in vivo animal models. Flow cytometry was used to describe the accumulation of submicron polymeric microcapsules in cellular spheroids, which allowed us to identify the main aspects of drug carrier-cell interactions within the spheroid. Spheroids were generated using 4T1 mouse breast cancer cells and healthy L929 mouse fibroblast cells. The arrangement of microcapsules with 300, 500, 1000 nm diameter with biocompatible shells in cell spheroids was evaluated in their intercellular and intracellular spaces.