Three-dimensional Tissue Models Research Articles

SLC37A4, gene responsible for glycogen storage disease type 1b, regulates gingival epithelial barrier function via JAM1 expression

Solute carrier family 37 member 4 (SLC37A4) is known to regulate glucose-6-phosphate transport from cytoplasm to the lumen of the endoplasmic reticulum, which serves to maintain glucose homeostasis. Glycogen storage disease type 1b (GSD1b) is caused by a mutation of SLC37A4, leading to a glycogenolysis defect. Although GSD1b cases are known to be complicated by periodontitis, the etiological molecular basis remains unclear. The present study investigated the effects of SLC37A4 on gingival barrier function. Examinations of immortalized human gingival epithelial (IHGE) cells showed SLC37A4 localized in the endoplasmic reticulum. SLC37A4 knockout decreased expression of JAM1, a tight junction-related protein, in IHGE cells. Using in silico analysis to investigate potential transcription factor binding sites, H6 family homeobox 3 (HMX3) was shown to be related to JAM1 expression. In HMX3-knockdown IHGE cells, JAM1 expression was markedly suppressed. Furthermore, HMX3 was scarcely detected in SLC37A4-knockout cells, while HMX3 overexpression restored JAM1 expression in those cells. Finally, using a three-dimensional multilayered gingival epithelial tissue model, knockout of SLC37A4 was also found to increase permeability to lipopolysaccharide and peptidoglycan, which was dependent on JAM1 expression. Specific downregulation of HMX3 by SLC37A4 and the consequent decrease in JAM1 expression provides findings indicating a molecular basis for the reduction in barrier function of gingival epithelial tissues in GSD1b cases.

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.

Human Cardiac Organoids: Quantification and Qualification in Cardiovascular Studies

Review Human Cardiac Organoids: Quantification and Qualification in Cardiovascular Studies Yingjuan Liu, Sabu Abraham, and Honglin Xu * Division of Cardiovascular Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, UK. * Correspondence: honglin.xu@postgrad.manchester.ac.uk Received: 4 April 2024; Revised: 29 May 2024; Accepted: 29 May 2024; Published: 29 August 2024 Abstract: The human cardiac organoids (hCOs) represent a three-dimensional (3D) tissue model that mirrors in vivo cardiac conditions. Recent advancements underscore the immense potential of hCOs in several areas including studying early cardiogenesis, modeling heart diseases, screening potential drugs, and even exploring possibilities for cardiac regeneration. Recognizing the pivotal role hCOs play across various applications, this review examines the evolution of key metrics and tools for assessing cardiac organoids tailored for diverse research objectives. Moreover, it deliberates on the limitations of cardiac organoids and outlines the prospective avenues for future research applications of hCOs.

Efficient fabrication of 3D bioprinted functional sensory neurons using an inducible Neurogenin-2 human pluripotent stem cell line

Three-dimensional (3D) tissue models have gained recognition for their improved ability to mimic the native cell microenvironment compared to traditional two-dimensional models. This progress has been driven by advances in tissue-engineering technologies such as 3D bioprinting, a promising method for fabricating biomimetic living tissues. While bioprinting has succeeded in generating various tissues to date, creating neural tissue models remains challenging. In this context, we present an accelerated approach to fabricate 3D sensory neuron (SN) structures using a transgenic human pluripotent stem cell (hPSC)-line that contains an inducible Neurogenin-2 (NGN2) expression cassette. The NGN2 hPSC line was first differentiated to neural crest cell (NCC) progenitors, then incorporated into a cytocompatible gelatin methacryloyl-based bioink for 3D bioprinting. Upregulated NGN2 expression in the bioprinted NCCs resulted in induced SN (iSN) populations that exhibited specific cell markers, with 3D analysis revealing widespread neurite outgrowth through the scaffold volume. Calcium imaging demonstrated functional activity of iSNs, including membrane excitability properties and voltage-gated sodium channel (NaV) activity. This efficient approach to generate 3D bioprinted iSN structures streamlines the development of neural tissue models, useful for the study of neurodevelopment and disease states and offering translational potential.

Promoting biological similarity by collagen microfibers in 3D colorectal cancer-stromal tissue: Replicating mechanical properties and cancer stem cell markers

The extracellular matrix (ECM) of cancer tissues is rich in dense collagen, contributing to the stiffening of these tissues. Increased stiffness has been reported to promote cancer cell proliferation, invasion, metastasis, and prevent drug delivery. Replicating the structure and mechanical properties of cancer tissue in vitro is essential for developing cancer treatment drugs that target these properties. In this study, we recreated specific characteristics of cancer tissue, such as collagen density and high elastic modulus, using a colorectal cancer cell line as a model. Using our original material, collagen microfibers (CMFs), and a constructed three-dimensional (3D) cancer-stromal tissue model, we successfully reproduced an ECM highly similar to in vivo conditions. Furthermore, our research demonstrated that cancer stem cell markers expressed in the 3D cancer-stromal tissue model more closely mimic in vivo conditions than traditional two-dimensional cell cultures. We also found that CMFs might affect an impact on how cancer cells express these markers. Our 3D CMF-based model holds promise for enhancing our understanding of colorectal cancer and advancing therapeutic approaches. Statement of significanceReproducing the collagen content and stiffness of cancer tissue is crucial in comprehending the properties of cancer and advancing anticancer drug development. Nonetheless, the use of collagen as a scaffold material has posed challenges due to its poor solubility, hindering the replication of a cancer microenvironment. In this study, we have successfully recreated cancer tissue-specific characteristics such as collagen density, stiffness, and the expression of cancer stem cell markers in three-dimensional (3D) colorectal cancer stromal tissue, utilizing a proprietary material known as collagen microfiber (CMF). CMF proves to be an ideal scaffold material for replicating cancer stromal tissue, and these 3D tissues constructed with CMFs hold promise in contributing to our understanding of cancer and the development of therapeutic drugs.

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.

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.

The influence of microwave ablation parameters on the positioning of trocar in different cancerous tissues: a numerical study

ABSTRACT The present study analyzed the microwave ablation of cancerous tumors located in six major cancer-prone organs and estimated the significance of input power and treatment time parameters in the apt positioning of the trocar into the tissue during microwave ablation. The present study has considered a three-dimensional two-compartment tumour-embedded tissue model. FEA based COMSOL Multiphysics software with inbuilt bioheat transfer, electromagnetic waves, heat transfer in solids and fluids, and laminar flow physics has been used to obtain the numerical results. Based on the mortality rates caused by cancer, the present study has considered six major organs affected by cancer, viz. lung, breast, stomach/gastric, liver, liver (with colon metastasis), and kidney for MWA analysis. The input power (100 W) and ablation times (4 minutes) with apt and inapt positioning of the trocar have been considered to compare the ablation volume of various cancerous tissues. The present study addresses one of the major problems clinicians face, i.e. the proper placement of the trocar due to poor imaging techniques and human error, resulting in incomplete tumor ablation and increased surgical procedures. The highest values of the ablation region have been observed for the liver, colon metastatic liver and breast cancerous tissues compared with other organs at the same operating conditions.

Abstract 221: Novel organotypic patient derived primary tumor tissues for oncology drug safety and efficacy studies

Abstract Organotypic, three-dimensional (3D) cancer tissue models (OCTM) have enabled investigators to develop predictive markers that can be used in preclinical cancer drug development. However, a drawback of the existing models is that they are mostly based on cell lines based are not physiological. In vitro development of 3D tumoroids using patient derived primary tumors (PDPT) in tumor microenvironment (TME) will have clinical significant and can predict human responses to cancer therapeutics. Here, we describe the generation of OCTM from patient-derived metastatic colorectal cancer, the second most deadly cancer. PDPT were grown in Matrigel for 5 passages and for biobanking purposes tumoroids were further expanded in faCellitate 384 well plates or Sun Bioscience plates to harvest large amounts of well-defined tumoroids cultured for over 21 days. Expanded tumoroids were collected and cryopreserved or gently mixed several times using a pipet tip to break up tumoroids into smaller fragments. These tumor fragments were mixed with endothelial cells, dendritic cells, fibroblasts and embedded in a collagen matrix to simulate TME. The collagen-cell mixture was then added to Transwells and induced to gel to form a collagen gel matrix that simulates TME. The tumoroids were cultured with a specialized medium that supports the growth of complex cell types. The tumor development was monitored daily using bright field microscopy and image analysis was used to monitor tumor size. To evaluate the utility of the OCTM, treatment was initiated at day 10 of the culture period by exposing tissues with the chemotherapeutic drug Cisplatin on days 0, 2, 4, 7, 9, and 11. After each treatment, OCTM tissues (N=2) were fixed for H&E and live/dead staining. Overall, the results of the study showed that: 1) PDPT can be expanded for 5 passages (this low passage number is not expected to change the phenotype of the tumor) and further expanded in high throughput format, 384 well plates, 2) the PDPT could be cultured with multiple cell types in a collagen-gel matrix to recreate the TME, 3) histological evaluation of the OCTM showed a glandular-like tumoroid, 4) immunohistochemical staining revealed that the PDPT in the in vivo-like microenvironment were positive for CK20, and 5) the PDPT shrank following 3 treatment cycles with Cisplatin (day 4). Conclusion: The data shows metastatic colorectal PDPT can be expanded in vitro and cultured in a complex tumor microenvironment on tissue culture inserts. These OCTMs can be used to screen drug safety and efficacy. This study opens new avenues for high-throughput cancer drug screening format using PDPT embedded in a in vivo like TME. This model can be used as a tool for personalized oncology drug testing and for translational assays tailored to other cancer types from our patient derived primary tumor pool. Citation Format: Seyoum Ayehunie, Megan Groves, Bryda Bryda, Alex Armento, Anthony Tolcher. Novel organotypic patient derived primary tumor tissues for oncology drug safety and efficacy studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 221.

Bioprinting with adipose stem cells and hydrogel modified with bioactive glass

Bioprinting research is focused on utilizing growth factors and multiple cell types to create clinically relevant three-dimensional (3D) tissue models using hydrogels. Rheological and biological challenges are two main factors that limit the creation of extrudable bioactive hydrogels. In this study, we investigate incorporation of fast dissolving and bioactive borate glass in different weight to volume percentages (0.075 to 0.6%) to alginate-gelatin (1:1) hydrogel to improve rheological properties and enable bioprinting with bioactive glass. The addition of glass improved the stiffness of the hydrogel. Human adipose-derived mesenchymal stem cells (ASCs) were uniformly mixed in this bioink at 1 × 106 cells/mL concentration, and spheroid specimens were cultured in both static and dynamic culture conditions. Grid-shaped scaffolds measuring ~18 × 18 × 1 mm3 were fabricated with the viable glass concentrations, and ASC viability was evaluated using Live/Dead assay. Despite immediate toxicity, an increased viability after 7 days with 0.15 w/v % or less borate glass content demonstrated the potential in utilizing highly resorbable calcium-releasing biomaterials such as bioactive glasses to modify hydrogels suitable for bioprinting cellularized 3D structures.

Multiscale transport and 4D time-lapse imaging in precision-cut liver slices (PCLS).

Monitoring cellular processes across different levels of complexity, from the cellular to the tissue scale, is important for understanding tissue structure and function. However, it is challenging to monitor and estimate these structural and dynamic interactions within three-dimensional (3D) tissue models. The aim of this study was to design a method for imaging, tracking, and quantifying 3D changes in cell morphology (shape and size) within liver tissue, specifically a precision-cut liver slice (PCLS). A PCLS is a 3D model of the liver that allows the study of the structure and function of liver cells in their native microenvironment. Here, we present a method for imaging liver tissue during anisosmotic exposure in a multispectral four-dimensional manner. Three metrics of tissue morphology were measured to quantify the effects of osmotic stress on liver tissue. We estimated the changes in the volume of whole precision cut liver slices, quantified the changes in nuclei position, and calculated the changes in volumetric responses of tissue-embedded cells. During equilibration with cell-membrane-permeating and non-permeating solutes, the whole tissue experiences shrinkage and expansion. As nuclei showed a change in position and directional displacement under osmotic stress, we demonstrate that nuclei could be used as a probe to measure local osmotic and mechanical stress. Moreover, we demonstrate that cells change their volume within tissue slices as a result of osmotic perturbation and that this change in volume is dependent on the position of the cell within the tissue and the duration of the exposure. The results of this study have implications for a better understanding of multiscale transport, mechanobiology, and triggered biological responses within complex biological structures.

Microvascular network based on the Hilbert curve for nutrient transport in thick tissue.

To address the uneven nutrient distribution within three-dimensional (3D) tissue models and organoids currently used in medical research, this study introduces a microvascular network based on the Hilbert curve. Our aim was to develop innovative solutions for enhancing nutrient supply in thick tissue models in vitro. By using 3D bioprinting, we engineered microvascular networks of varying Hilbert orders and validated their efficacy in enhancing nutrient uniformity through numerical simulations and experiments. These networks facilitated broader and more uniform nutrient distribution throughout the thick tissue models, particularly the 2° Hilbert microvascular structure, which occupies less space and significantly reduces regions of cellular death. Furthermore, we explored the potential of assembling larger tissue constructs using the 2° Hilbert microvascular network, showcasing its applicability in constructing large-scale biological models. The findings suggest that the 2° Hilbert microvascular structure is particularly effective in ensuring adequate nutrient delivery, thus enhancing the viability and functionality of large-volume tissue models. These innovations hold significant promise for advancing the fields of tissue engineering and regenerative medicine by improving nutrient delivery to in vitro thick tissue block models. This provides a robust foundation for future in vitro research and clinical applications, potentially leading to more effective treatments and interventions in the medical field. The development of these microvascular networks represents a crucial step forward in overcoming the limitations of current 3D tissue models and organoids, paving the way for more sophisticated and reliable biomedical research tools.

Role of Hepatic Stellate and Liver Sinusoidal Endothelial Cells in a Human Primary Cell Three-Dimensional Model of Nonalcoholic Steatohepatitis

Nonalcoholic steatohepatitis (NASH) is an inflammatory and fibrotic liver disease that has reached epidemic proportions and has no approved pharmacologic therapies. Research and drug development efforts are hampered by inadequate preclinical models. This research describes a three-dimensional bioprinted liver tissue model of NASH built using primary human hepatocytes and nonparenchymal liver cells (hepatic stellate cells, liver sinusoidal endothelial cells, and Kupffer cells) from either healthy or NASH donors. Three-dimensional tissues bioprinted with cells sourced from diseased patients showed a NASH phenotype, including fibrosis. More importantly, this NASH phenotype occurred without the addition of disease-inducing agents. Bioprinted tissues composed entirely of healthy cells exhibited significantly less evidence of disease. The role of individual cell types in driving the NASH phenotype was examined by producing chimeric bioprinted tissues composed of healthy cells together with the addition of one or more diseased nonparenchymal cell types. These experiments reveal a role for both hepatic stellate and liver sinusoidal endothelial cells in the disease process. This model represents a fully human system with potential to detect clinically active targets and eventually therapies.

Nano- and Microstructures of Collagen-Nanocellulose Hydrogels as Engineered Extracellular Matrices.

The extracellular matrix (ECM) is the fundamental acellular element of human tissues, providing their mechanical structure while delivering biomechanical and biochemical signals to cells. Three-dimensional (3D) tissue models commonly use hydrogels to recreate the ECM in vitro and support the growth of cells as organoids and spheroids. Collagen-nanocellulose (COL-NC) hydrogels rely on the blending of both polymers to design matrices with tailorable physical properties. Despite the promising application of these biomaterials in 3D tissue models, the architecture and network organization of COL-NC remain unclear. Here, we investigate the structural effects of incorporating NC fibers into COL hydrogels by small-angle neutron scattering (SANS) and ultra-SANS (USANS). The critical hierarchical structure parameters of fiber dimensions, interfiber distance, and coassembled open structures of NC and COL in the absence and presence of cells were determined. We found that NC expanded and increased the homogeneity in the COL network without affecting the inherent fiber properties of both polymers. Cells cultured as spheroids in COL-NC remodeled the hydrogel network without a significant impact on its architecture. Our study reveals the polymer organization of COL-NC hydrogels and demonstrates SANS and USANS as exceptional techniques to reveal nano- and micron-scale details on polymer organization, which leads to a better understanding of the structural properties of hydrogels to engineer novel ECMs.

Patchy fibrosis promotes trigger-substrate interactions that both generate and maintain atrial fibrillation.

Fibrosis has been mechanistically linked to arrhythmogenesis in multiple cardiovascular conditions, including atrial fibrillation (AF). Previous studies have demonstrated that fibrosis can create functional barriers to conduction which may promote excitation wavebreak and the generation of re-entry, while also acting to pin re-entrant excitation in stable rotors during AF. However, few studies have investigated the role of fibrosis in the generation of AF triggers in detail. We apply our in-house computational framework to study the impact of fibrosis on the generation of AF triggers and trigger-substrate interactions in two- and three-dimensional atrial tissue models. Our models include a reduced and efficient description of stochastic, spontaneous cellular triggers as well as a simple model of heterogeneous inter-cellular coupling. Our results demonstrate that fibrosis promotes the emergence of focal excitations, primarily through reducing the electrotonic load on individual fibre strands. This enables excitation to robustly initiate within these single strands before spreading to neighbouring strands and inducing a full tissue focal excitation. Enhanced conduction block can allow trigger-substrate interactions that result in the emergence of complex, re-entrant excitation patterns. This study provides new insight into the mechanisms by which fibrosis promotes the triggers and substrate necessary to induce and sustain arrhythmia.

Organotypic model of the gingiva for studying bacterial and viral pathogens implicated in periodontitis

ABSTRACT Background Three-dimensional (3D) tissue models bridge the gap between conventional two-dimensional cell cultures and animal models. The aim of this study was to develop an organotypic 3D gingival (OTG) model to provide a tool to investigate bacterial and viral pathogens in periodontitis. Methods The OTG model composed of gingival fibroblasts (GFs) and telomerase-immortalized gingival keratinocytes (TIGKs) was constructed and applied to study infections by Porphyromonas gingivalis and herpes simplex virus 1 (HSV-1). Immunohistochemical staining, confocal microscopy, qPCR, titration techniques, and colony-forming unit counts were applied to interrogate epithelial markers expression, monitor P. gingivalis and HSV-1 presence, and evaluate the immune response along with the efficiency of antimicrobial drugs. Results The OTG model resembled the morphology of the human gingiva. During infection, both pathogens penetrated deep into the tissue and persisted for a few days with P. gingivalis also forming a biofilm on the cell surface. The infection triggered the expression of inflammatory mediators in cells and both pathogens were efficiently eliminated by specific antimicrobials. Conclusions Presented OTG model constitutes a simple and convenient tool to study the interaction between bacterial and viral pathogens within the gingival tissue, including penetration, persistence and biofilm formation. It is also suitable to examine the efficiency of antimicrobial drugs.

In-Vitro Decidualization With Different Progesterone Concentration: Development of a Hormone-Responsive 3D Endometrial Tissue Using Rat Endometrial Tissues.

Infertility in women is associated with various uterine and ovarian disorders. Treatment strategies for infertility can range from medications to embryo implantation through assisted reproductive technology (ART). ART has enabled considerable progress; however, there is currently no treatment to replacethe endometrium itself. Decidualization requires a complex interaction between endometrial tissue and estrogen and progesterone.We aimed to createa three-dimensional endometrial-like tissue model usingin-vitro cell sheet engineering with rat endometrium, and culturecells at different progesterone concentrations to mimic local concentrations.Histological and morphological changes revealed that development of the endometrial-like tissuewas not proportional to progesterone concentrations in terms of thickness, number of endometrial glands, or area fraction of intimal glands. These results suggest that decidualization may not be commensurate with the local endometrial progesterone concentration.Notably, the number of endometrial glands increased in the high concentration group and compaction occurred, indicating thatthe endometrial conditions in the high concentration group may be most conducive to increase pregnancy rates. Thesefindings suggest that there may be an "optimal progesterone concentration" for decidualization, application of which may lead to new strategies for improving pregnancy rates in women with infertility.

A quantitative morphospace of multicellular organ design in the plant Arabidopsis

Organ function emerges from the interactions between their constituent cells. The investigation of cellular organization can provide insight into organ function following structure-function relationships. Here, we investigate the extent to which properties in cellular organization can arise "for free" as an emergent property of embedding cells in space versus those that are actively generated by patterning processes. Default cellular configurations were established using three-dimensional (3D) digital tissue models. Network-based analysis of these synthetic cellular assemblies established a quantitative topological baseline of cellular organization, granted by virtue of passive spatial packing and the minimal amount of order that emerges for free in tessellated tissues. A 3D cellular-resolution digital tissue atlas for the model plant species Arabidopsis was generated, and the extent to which the organs in this organism conform to the default configurations was established through statistical comparisons with digital tissue models. Cells in different tissues of Arabidopsis do not conform to random packing arrangements to varying degrees. Most closely matching the random models was the undifferentiated shoot apical meristem (SAM) from which aerial organs emanate. By contrast, leaf and sepal tissue showed the greatest deviation from this baseline, suggesting these to be the most "complex" tissues in Arabidopsis. Investigation of the patterning principles responsible for the gap between these tissues and default patterns revealed cell elongation and the introduction of air spaces to contribute toward additional organ patterning complexity. This work establishes a quantitative morphospace to understand the principles of organ construction and its diversity within a single organism.

Durable 3D murine ex vivo retina glaucoma models for optical coherence tomography.

Durable and standardized phantoms with optical properties similar to native healthy and disease-like biological tissues are essential tools for the development, performance testing, calibration and comparison of label-free high-resolution optical coherence tomography (HR-OCT) systems. Available phantoms are based on artificial materials and reflect thus only partially ocular properties. To address this limitation, we have performed investigations on the establishment of durable tissue phantoms from ex vivo mouse retina for enhanced reproduction of in vivo structure and complexity. In a proof-of-concept study, we explored the establishment of durable 3D models from dissected mouse eyes that reproduce the properties of normal retina structures and tissue with glaucoma-like layer thickness alterations. We explored different sectioning and preparation procedures for embedding normal and N-methyl-D-aspartate (NMDA)-treated mouse retina in transparent gel matrices and epoxy resins, to generate durable three-dimensional tissue models. Sample quality and reproducibility were quantified by thickness determination of the generated layered structures utilizing computer-assisted segmentation of OCT B-scans that were acquired with a commercial HR-OCT system at a central wavelength of 905 nm and analyzed with custom build software. Our results show that the generated 3D models feature thin biological layers close to current OCT resolution limits and glaucoma-like tissue alterations that are suitable for reliable HR-OCT performance characterization. The comparison of data from resin-embedded tissue with native murine retina in gels demonstrates that by utilization of appropriate preparation protocols, highly stable samples with layered structures equivalent to native tissues can be fabricated. The experimental data demonstrate our concept as a promising approach toward the fabrication of durable biological 3D models suitable for high-resolution OCT system performance characterization supporting the development of optimized instruments for ophthalmology applications.

Cigarette smoke extract impairs gingival epithelial barrier function

We previously showed that junctional adhesion molecule 1 (JAM1) and coxsackievirus and adenovirus receptor (CXADR), tight junction-associated proteins, have important roles to maintain epithelial barrier function in gingival tissues. Smoking is considered to be a significant risk factor for periodontal disease. The present study was conducted to examine the effects of cigarette smoke extract (CSE) on JAM1 and CXADR in human gingival epithelial cells. CSE was found to cause translocation of JAM1 from the cellular surface to EGFR-positive endosomes, whereas CXADR did not. Using a three-dimensional multilayered gingival epithelial tissue model, CSE administration was found to increase permeability to lipopolysaccharide and peptidoglycan, whereas overexpression of JAM1 in the tissue model prevented penetration by those substrates. Furthermore, vitamin C increased JAM1 expression, and inhibited penetration of LPS and PGN induced by CSE. These findings strongly suggest that CSE disrupts gingival barrier function via dislocation of JAM1, thus allowing bacterial virulence factors to penetrate into subepithelial tissues. Furthermore, they indicate that vitamin C increases JAM1 expression and prevents disruption of gingival barrier function by CSE.