In vitro models最新文献

Purpose: For optimization of respiratory drug delivery, the selection of suitable in vitro cell models plays an important role in predicting the efficacy and safety of (bio)pharmaceutics and pharmaceutical formulations. Therefore, an in-depth comparison of different primary and permanent in vitro cellular airway models was performed with a focus on selecting a suitable model for inhalative antibodies.

Methods: Primary cells isolated from the porcine trachea were compared with the established human cell lines CaLu3 and RPMI 2650. The in vitro models were characterized for different epithelial markers by real-time quantitative polymerase chain reaction, which provides insight into the cellular composition of each model. For a few selected markers, the results from RT-qPCR were confirmed via immunofluorescence. Barrier integrity was assessed by transepithelial electrical resistance measurements and FITC-dextran permeability.

Results: Primary cell models retain key features of the respiratory epithelium, e.g., the formation of a tight epithelial barrier, mucin production, and the presence of club/basal cells. Furthermore, the expression of Fc receptors in the primary cell models closely resembles that in respiratory mucosal tissue, an essential parameter to consider when developing therapeutic antibodies for inhalation.

Conclusion: The study underlines the importance of selecting wisely appropriate in vitro models. Despite the greater effort and variability in cultivating primary airway cells, they are far superior to permanent cells and a suitable model for drug development.

Supplementary information: The online version contains supplementary material available at 10.1007/s44164-024-00079-y.

Background: Extracellular matrix (ECM) is a three-dimensional (3D) structure found around cells in the tissues of many organisms. It is composed mainly of fibrous proteins, such as collagen and elastin, and adhesive glycoproteins, such as fibronectin and laminin-as well as proteoglycans, such as hyaluronic acid. The ECM performs several essential functions, including structural support of tissues, regulation of cell communication, adhesion, migration, and differentiation by providing biochemical and biomechanical cues to the cells. Pancreatic β-cells have been previously shown to be responsive to the surrounding mechanical stress, impacting their insulin-secreting function.

Purpose: We aimed to derive a physiologically relevant in vitro model of pancreatic tissue by using an innovative synthesised porous ECM that mimics the native tissue microenvironment and mechanical properties.

Methods: Here we performed mechanical, physico-chemical and functional characterisation of a synthetic hydrogel ECM, composed of hyaluronic acid cross-linked with collagen types I and VI and modified with fibronectin. The hydrogel was used as a 3D cell culture scaffold for the MIN6 insulinoma cell line. Cell proliferation, viability, gene expression, and insulin secretion in response to glucose stimulus were assessed and contrasted with classic monolayer culture.

Results: The biomaterial exhibited a shear modulus of 815.37 kPa and a distinctive viscoelastic response. MIN6 cells showed a higher proliferation and viability rates and maintained insulin secretion in response to glucose stimulus and β-cell identity gene expression when cultured in the 3D hydrogel compared to monolayer culture.

Conclusion: Our study demonstrated the potential of this biomimetic hydrogel scaffold as an innovative matrix enabling better in vitro models to study disease physiopathology.

Supplementary information: The online version contains supplementary material available at 10.1007/s44164-024-00078-z.

Obesity is associated with several comorbidities that cause high mortality rates worldwide. Thus, the study of adipose tissue (AT) has become a target of high interest because of its crucial contribution to many metabolic diseases and metabolizing potential. However, many AT-related physiological, pathophysiological, and toxicological mechanisms in humans are still poorly understood, mainly due to the use of non-human animal models. Organ-on-chip (OoC) platform is a promising alternative to animal models. However, the use of adipose-derived human mesenchymal stem cells (hASCs) in these models is still scarce, and more knowledge on the effects properties of culturing hASCs in OoC models is needed. Here, we present the development of an OoC using hASCs to assess adipogenic differentiation. The device capability to increase hASC differentiation levels was confirmed by Nile red staining to verify lipid droplets inside cells after 10 days of culture and fluid flow of 10 µL/h. The Adipo-on-a-chip system increases hASC proliferation and differentiation area compared with the standard culture method under static conditions (96-well plates) verified in hASCs from different donors by image analysis of cells stained with Nile red. The expression of the gene FABP4 is lower in the MPS, which calls attention to different homeostasis and control of lipids in cells in the MPS, compared with the plates. An increase of hASC proliferation in the MPS related to the 96-well plate was verified through protein Ki-67 expression. Cell and nuclei morphology (area, roundness, perimeter, width, length, width to length rate, symmetry, compactness, axial and radial properties to nuclei, and texture) and dominant direction of cells inside the MPS were evaluated to characterize hASCs in the present model. The presented microphysiological system (MPS) provides a promising tool for applications in mechanistic research aiming to investigate adipogenesis in AT and toxicological assessment based on the hASC differentiation potential.

Zinc (Zn) and its alloys have been the focus of recent materials and manufacturing research for orthopaedic implants due to their favorable characteristics including desirable mechanical strength, biodegradability, and biocompatibility. In this research, a novel process involving additive manufacturing (AM) augmented casting was employed to fabricate zinc-magnesium (Zn-0.8 Mg) artifacts with surface lattices composed of triply periodic minimal surfaces (TPMS), specifically gyroid. Scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS) analysis confirmed that Zn-Mg intermetallic phases formed at the grain boundary. Micro indentation testing resulted in hardness value ranging from 83.772 to 99.112 HV and an elastic modulus varying from 92.601 to 94.625 GPa. Results from in vitro cell culture experiments showed that cells robustly survived on both TPMS and solid scaffolds, confirming the suitability of the material and structure as biomedical implants. This work suggests that this novel hybrid manufacturing process may be a viable approach to fabricating next generation biodegradable orthopaedic implants.

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Owing to increased pressure from ethical groups and the public to avoid unnecessary animal testing, the need for new, responsive and biologically relevant in vitro models has surged. Models of the human alveolar epithelium are of particular interest since thorough investigations into air pollution and the effects of inhaled nanoparticles and e-cigarettes are needed. The lung is a crucial organ of interest due to potential exposures to endogenous material during occupational and ambient settings. Here, an in vitro model of the alveolar barrier has been created in preparation for use in the quasi-air liquid interface (qALI) and (aerosol) air-liquid interface (ALI) exposures. The model consists of an alveolar type 1-like cell line (TT1), an alveolar type 2-like cell line (NCI-H441) and a model of (alveolar) macrophages (dTHP-1). The model formulates a complex, multi-cellular system, cultured at the air-liquid interface, that mimics the apical layer of the alveolar epithelial region in the human lung. Characterisation data has shown that both TT1 and NCI-H441 epithelial cells are able to be cultured together in addition to dTHP-1 cells through imaging (morphology), pro-inflammatory response and viability measurements. This dataset also demonstrates evidence of a reasonable barrier created by the cell culture in comparison to negative controls. Furthermore, it shows that while maintaining a low baseline of (pro)-inflammatory mediator expression during normal conditions, the model is highly responsive to inflammatory stimuli. This model is proposed to be suitable for use in toxicology testing of inhaled exogenous agents.

Supplementary information: The online version contains supplementary material available at 10.1007/s44164-024-00075-2.

Wound debridement is commonplace in expediting wound healing in the clinic. Despite this, there are limited resources available for simulation training for practitioners prior to facing real-life patients. Typically, citrus peels or porcine skin are employed in a vain attempt to improve debridement proficiency, yet these fail to provide a realistic experience of the textures and consistencies of wounds. Therefore, there is a clear unmet need for a safe and effective tool that can facilitate hands-on learning under the instruction of an experienced debrider. To fill this niche, a life-like wound model was designed and developed, featuring leathery necrotic eschar, an intermediary sloughy layer, and a rough granulation tissue in the bottommost layer, all within a healthy skin base. The healthy tissue portion of the model was designed to exhibit similar mechanical properties to those found in human skin. Likewise, the sloughy layer was viscous enough to remain within the model under static conditions yet could be removed using any appropriate debriding tool synonymous with real slough. Mechanical testing of the necrotic eschar revealed brittle fracture behaviour, akin to what is observed in patients. Each layer of the wound model provided the visual and haptic feedback of how it would look and feel to debride a patient's wound in the clinic, giving invaluable experience for potential trainees in a safe and effectual way. It is envisaged that these models can be developed in a personalised way to suit the individual needs of the user, such as incorporating underlying models of bone or tendon, while retaining the key elements of the wound, which make it successful. This model is proposed as an important step forward in bridging the gap between becoming a newly qualified debriding practitioner and encountering the first wound in the clinic, subsequently improving the confidence of the debrider and enhancing patient outcomes.

Supplementary information: The online version contains supplementary material available at 10.1007/s44164-024-00071-6.

[This corrects the article DOI: 10.1007/s44164-023-00055-y.].