Nanoparticles allow for controlled and targeted drug delivery to diseased tissues and therefore bypass systemic side effects. Spatiotemporal control of drug release can be achieved by nanocarriers ...that respond to elevated levels of disease-specific enzymes. For example, matrix metalloproteinase 9 (MMP9) is overexpressed in tumors, is known to enhance the metastatic potency of malignant cells, and has been associated with poor prognosis of lung cancer. Here, we report the synthesis of mesoporous silica nanoparticles (MSNs) tightly capped by avidin molecules via MMP9 sequence-specific linkers to allow for site-selective drug delivery in high-expressing MMP9 tumor areas. We provide proof-of-concept evidence for successful MMP9-triggered drug release from MSNs in human tumor cells and in mouse and human lung tumors using the novel technology of ex vivo 3D lung tissue cultures. This technique allows for translational testing of drug delivery strategies in diseased mouse and human tissue. Using this method we show MMP9-mediated release of cisplatin, which induced apoptotic cell death only in lung tumor regions of Kras mutant mice, without causing toxicity in tumor-free areas or in healthy mice. The MMP9-responsive nanoparticles also allowed for effective combinatorial drug delivery of cisplatin and proteasome inhibitor bortezomib, which had a synergistic effect on the (therapeutic) efficiency. Importantly, we demonstrate the feasibility of MMP9-controlled drug release in human lung tumors.
In our previous study, we produced a microfluidic device (MFD) which successfully maintained spermatogenesis for over 6 months in mouse testis tissues loaded in the device. In the present study, we ...developed a new MFD, a monolayer device (ML-D) with a barrier structure consisting of pillars and slits, which is simpler in design and easier to make. This ML-D was also effective for inducing mouse spermatogenesis and maintained it for a longer period than the conventional culture method. In addition, we devised a way of introducing sample tissue into the device during its production, just before bonding the upper layer of polydimethylsiloxane (PDMS) and bottom glass slide. The tissue can obtain nutrients horizontally from the medium running beside it and oxygen vertically from above through PDMS. In addition, the glass slide set at the bottom improved the visibility of the sample tissue with an inverted microscope. When we took photos of cultured tissue of the Acr-Gfp transgenic mouse testis in ML-D sequentially every day, morphological changes of the acrosome during spermiogenesis were successfully recorded. The ML-D is simple in design and useful for culturing testis tissue for inducing and maintaining spermatogenesis with clearer visibility. Due to the new method of sample loading, tissues other than testis should also be applicable.
Abstract New molecular diagnostic tools have recently allowed the discovery of human rhinovirus species C (HRV-C) that may be overrepresented in children with lower respiratory tract complications. ...Unlike HRV-A and HRV-B, HRV-C cannot be propagated in conventional immortalized cell lines and their biological properties have been difficult to study. Recent studies have described the successful amplification of HRV-C15, HRV-C11, and HRV-C41 in sinus mucosal organ cultures and in fully differentiated human airway epithelial cells. Consistent with these studies, we report that a panel of clinical HRV-C specimens including HRV-C2, HRV-C7, HRV-C12, HRV-C15, and HRV-C29 types were all capable of mediating productive infection in reconstituted 3D human primary upper airway epithelial tissues and that the virions enter and exit preferentially through the apical surface. Similar to HRV-A and HRV-B, our data support the acid sensitivity of HRV-C. We observed also that the optimum temperature requirement during HRV-C growth may be type-dependent.
Techniques for tissue culture have seen significant advances during the last decades and novel 3D cell culture systems have become available. To control their high complexity, experimental techniques ...and their Digital Twins (modelling and computational tools) are combined to link different variables to process conditions and critical process parameters. This allows a rapid evaluation of the expected product quality. However, the use of mathematical simulation and Digital Twins is critically dependent on the precise description of the problem and correct input parameters. Errors here can lead to dramatically wrong conclusions. The intention of this review is to provide an overview of the state‐of‐the‐art and remaining challenges with respect to generating input values for computational analysis of mass and momentum transport processes within tissue cultures. It gives an overview on relevant aspects of transport processes in tissue cultures as well as modelling and computational tools to tackle these problems. Further focus is on techniques used for the determination of cell‐specific parameters and characterization of culture systems, including sensors for on‐line determination of relevant parameters. In conclusion, tissue culture techniques are well‐established, and modelling tools are technically mature. New sensor technologies are on the way, especially for organ chips. The greatest remaining challenge seems to be the proper addressing and handling of input parameters required for mathematical models. Following Good Modelling Practice approaches when setting up and validating computational models is, therefore, essential to get to better estimations of the interesting complex processes inside organotypic tissue cultures in the future.
Salvia miltiorrhiza Bunge (Lamiaceae) root, generally called Danshen, is an important herb in Chinese medicine widely used for treatment of cardiovascular diseases. Diterpenoid tanshinons are major ...bioactive constituents of Danshen with notable pharmacological activities and the potential as new drug candidates against some important human diseases. The importance of Danshen for traditional and modern medicines has motivated the research interest over two decades in the biosynthesis and biotechnological production of tanshinones. Although diterpenes in plants are presumably derived from the non-mevalonate (MVA) pathway, tanshinone biosynthesis in S. miltiorrhiza may also depend on the MVA pathway based on some key enzymes and genes detected in the early steps of these pathways. Plant tissue cultures are the major biotechnological processes for rapid production of tanshinones and other bioactive compounds in the herb. Various in vitro cultures of S. miltiorrhiza have been established, including cell suspension, adventitious root, and hairy root cultures, which can accumulate the major tanshinones as in the plant roots. Tanshinone production in cell and hairy root cultures has been dramatically enhanced with various strategies, including medium optimization, elicitor stimulation, and nutrient feeding operations. This review will summarize the above developments and also provide our views on future trends.
Exposure to air pollutants, such as particulate matter (PM), has been implicated in neurodegenerative disorders including Alzheimer's disease (AD). However, direct effects of PM on production of ...β-amyloid (Aβ), a key pathogenic molecule in AD, and its underlying mechanism are still elusive. Given PM's potential to induce oxidative stress in other tissues, we hypothesized that poly(ADP-ribose) polymerase (PARP-1) might be involved in PM-induced neurotoxicity. To address this, we used an ex vivo model of AD, the organotypic hippocampal slice tissue culture from old (12-14 months-of-age) triple transgenic 3xTg-AD mice. First, we observed that fine PM (aerodynamic diameter < 4 μm) can dose-dependently activate PARP-1 and decrease NAD+ levels in Neuro2A cells. PARP-1 activation did occur under concentrations of PM which did not affect cell viability. Next, we observed that direct treatment of PM increased Aβ levels and activated glial cells in the ex vivo hippocampal tissues of 3xTg-AD mice. PM-induced glial activation was most prominent in CA1 region of the hippocampal tissue. Notably, we found that pharmacological inhibition of PARP-1 reversed both PM-induced Aβ increase and glial activation, arguing the possible involvement of PARP-1 in PM-induced AD pathogenesis. Our findings suggest that PARP-1 might be a potential molecular target, responsible for mediating negative effects of PM on the brain. Modulating PARP-1 activity could be a promising approach to prevent or alleviate PM-related environmental neurotoxicity which could initiate AD pathogenesis.
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•Particulate matter (PM) activates PARP-1 in neuronal cells.•PM increases beta-amyloid in ex vivo hippocampus of 3xTg-AD mouse.•PM activates glial cells in ex vivo hippocampus of 3xTg-AD mouse.•Pharmacological inhibition of PARP-1 reverses PM-induced AD pathologies.•PARP-1 inhibition would be a promising approach counteracting PM's neurotoxicity.
This is a meditation on the role of absence during the COVID-19, especially the ways absences are felt and experienced. It explores the roles of bodies as both symbols and material. Bodies are both ...thought through the logic of borders and difference but also as the raw resources of scientific investigations. This is all examined within and against “education” both in my and in my students’ (pre and in-service teachers) classes and our anxieties of not knowing the what or how we of our jobs in these conditions.
Current systems for plant-based biomaterial production are inefficient and place unsustainable demands on environmental resources. This work proposes a novel solution to these shortcomings based on ...selective cultivation of tunable plant tissues using scalable, land-free techniques unconstrained by seasonality, climate, or local resource availability. By limiting biomass cultivation to only desirable plant tissues, ex planta farming promises to improve yields while reducing plant waste and competition for arable land. Employing a Zinnia elegans model system, this work provides the first proof-of-concept demonstration of isolated, tissue-like plant material production in vitro by way of gel-mediated cell culture. Parameters governing cell development and morphology including hormone concentrations, medium pH, and initial cell density are optimized and implemented to demonstrate the tunability of cultured biomaterials at cellular and macroscopic scales. Targeted deposition of cell-doped, nutrient-rich gel scaffolds via casting and 3D bioprinting enable biomaterial growth in near-final form, reducing downstream processing requirements. These investigations demonstrate the implementation of plant cell culture in a new application space, propose novel methods for quantification and evaluation of cell development, and characterize morphological developments in response to critical culture parameters—illustrating the feasibility and potential of the proposed techniques.
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•Agriculture-free growth of plant materials is feasible, controllable, limits waste.•Plant tissue-like materials are selectively grown free of unnecessary organs.•Adjusting scaffold properties tunes cellular make-up of grown material.•New metrics map cell growth and development to culture conditions, allow optimizing.•Bioprinting and injection molding are used to shape scaffold and grown material.
During the last decades, the study of cell behavior was largely accomplished in uncoated or extracellular matrix (ECM)-coated plastic dishes. To date, considerable cell biological efforts have tried ...to model in vitro the natural microenvironment found in vivo. For the lung, explants cultured ex vivo as lung tissue cultures (LTCs) provide a three-dimensional (3D) tissue model containing all cells in their natural microenvironment. Techniques for assessing the dynamic live interaction between ECM and cellular tissue components, however, are still missing. Here, we describe specific multidimensional immunolabeling of living 3D-LTCs, derived from healthy and fibrotic mouse lungs, as well as patient-derived 3D-LTCs, and concomitant real-time four-dimensional multichannel imaging thereof. This approach allowed the evaluation of dynamic interactions between mesenchymal cells and macrophages with their ECM. Furthermore, fibroblasts transiently expressing focal adhesions markers incorporated into the 3D-LTCs, paving new ways for studying the dynamic interaction between cellular adhesions and their natural-derived ECM. A novel protein transfer technology (FuseIt/Ibidi) shuttled fluorescently labeled α-smooth muscle actin antibodies into the native cells of living 3D-LTCs, enabling live monitoring of α-smooth muscle actin-positive stress fibers in native tissue myofibroblasts residing in fibrotic lesions of 3D-LTCs. Finally, this technique can be applied to healthy and diseased human lung tissue, as well as to adherent cells in conventional two-dimensional cell culture. This novel method will provide valuable new insights into the dynamics of ECM (patho)biology, studying in detail the interaction between ECM and cellular tissue components in their natural microenvironment.
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