Search Results (46)

Three-dimensional bioprinting integrating living cells and bioactive materials enables the fabrication of scaffold structures supporting diverse cellular growth and metabolism. Microalgae are among the most promising microbial platforms for the construction of photosynthetic cell factories, while the current industrial-scale cultivation of microalgae remains predominantly dependent on traditional liquid submerged systems, imposing limitations on commercial viability due to both process and economic constraints. Encapsulation of microalgae within bioactive matrices combined with 3D bioprinting to fabricate customized structures has been explored to address the limitations of submerged cultivation, which are expected to expand microalgal applications and establish new research directions in microalgal biotechnology. This review analyzes both matrices and methods of 3D bioprinting, summarizing the advancement of microalgae-based 3D bioprinting into six main domains including living building materials, biophotovoltaics, photosynthetic biosynthesis, bioremediation, tissue engineering, and food engineering. Lastly, synthetic biology-informed perspectives are provided on future developments of 3D bioprinting technologies and their potential in microalgal research. Full article

Approximately 20–30% of cultivated oyster mushrooms (Pleurotus ostreatus) are classified as low grade due to morphological and visual imperfections or mechanical damage, representing significant waste in mushroom production systems. This review examines the structural and biochemical properties of P. ostreatus, particularly focusing on cell wall components including chitin, β-glucans, and mannogalactans, which provide crucial rheological characteristics for 3D printing. The literature results demonstrate that these natural polysaccharides contribute essential viscosity, water-binding capacity, and mechanical stability required for printable edible inks. Notably, the mushroom stipe contains significantly higher concentrations of glucans compared to the cap, with 57% more α-glucans and 33% more β-glucans. The unique combination of rigidity from chitin, elasticity from β-glucans, and water retention capabilities creates printable structures that maintain shape fidelity while delivering nutritional benefits. This approach addresses dual challenges in sustainable food systems by reducing agricultural waste streams while advancing eco-friendly food innovation. The integration of mushroom-derived biomaterials into 3D printing technologies offers a promising pathway toward developing nutrient-rich, functional foods within a regenerative production model. Full article

This study proposes a continuous preparation strategy for poly (l-lactic acid) (PLLA) scaffolds with oriented hierarchical microporous structures for bone repair. A PLLA-oriented multi-stage microporous bone repair scaffold (hereafter referred to as the oriented multi-stage microporous scaffold) was designed using a novel extrusion foaming technology that integrates fused deposition modeling (FDM) 3D printing with supercritical carbon dioxide (SC-CO₂) microcellular foaming technology. The influence of the 3D-printed structure on the microcellular morphology of the oriented multi-stage microporous scaffold was investigated and optimized. The combination of FDM and SC-CO₂ foaming technology enables a continuous extrusion foaming process for preparing oriented multi-stage microporous scaffolds. The mechanical strength of the scaffold reached 15.27 MPa, meeting the requirements for bone repair in a low-load environment. Notably, the formation of open pores on the surface of the oriented multi-stage microporous scaffold positively affected cell proliferation, differentiation, and activity, as well as the expression of anti-inflammatory and pro-inflammatory factors. In vitro cell experiments (such as CCK-8) showed that the cell proliferation rate in the oriented multi-stage microporous scaffold reached 100–300% after many days of cultivation. This work provides a strategy for the design and manufacture of PLLA scaffolds with hierarchical microcellular structures and biocompatibility for bone repair. Full article

Virus-like particles (VLPs) have the potential to become a versatile carrier platform for vaccination against multiple diseases. In the light of short process development timelines and the demand for reliable and robust processes, metabolic modeling of cell culture processes offers great advantages when coupled with a Quality-by-Design (QbD) development approach. A previous work was able to demonstrate the accurate prediction of HEK293F PiggyBac cell concentration as well as VLP titer and metabolite production with a reduced metabolic model. This work presents the reduced metabolic model for a more productive cell line Sleeping Beauty and emphasizes the need for model re-parameterization when the producer cell line changes. The goal of precise prediction for a fed-batch and continuous HEK293 cultivation can, therefore, be achieved. In terms of decision-making for downstream unit operations, a soft sensor for the prediction of main impurities like proteins and DNA was introduced for the first time for the production of lentiviral vectors with several terms describing the release of impurities like DNA and proteins, growth-related protein production, and enzymatic degradation activity associated with cell dissociation in an accurate manner. The additional information can contribute to a more efficient design phase by reducing experimental effort as well as during cultivation with data-based decision-making. With the aid of real-time process data acquisition through process analytical technology (PAT), its predictive power can be enhanced and lead to more reliable processes. Full article

Cultivated meat, developed through cell culture technology, is emerging as a promising solution that closely mimics both the flavor and nutrient profiles of conventional meat. One key component that contributes to the flavor of meat is fat content. In this study, bovine adipose-derived stem cells (bADSCs) were cultured for the production of alternative fat in vitro. The expression of mesenchymal stem cell (MSC) markers (CD29, CD73, and CD105) and colony forming efficiency were assessed to characterize bADSCs. bADSCs were differentiated into adipocytes to produce cultivated fat in 2D or 3D culture. The cultivated fat was analyzed by gas chromatography to verify the similarity of the fatty acids of animal-derived fat. Our results show that bADSCs have characteristics of MSC and could differentiate into adipocyte. The ratio of unsaturated fatty acids and saturated fatty acids in cultivated fat and adipose tissue was similar. Adipogenic differentiation of ADSCs using a textured vegetable protein (TVP) scaffold could form the lipid droplets in the TVP. This study demonstrated the establishment of a culture system for the fat production from bADSCs in vitro. The fat produced through bADSCs shows the potential to be used in the composition of hybrid-cultivated meat. Full article

The Satsuma mandarin, a prominent fresh citrus variety cultivated in Asia, is susceptible to fruit cracking, a physiological disorder that significantly impacts yield and economic efficiency. This phenomenon occurs during the fruit expansion phase. The present study sought to further elucidate the correlation between citrus fruit cracking and fruit peel development or mineral elements, as well as to propose efficacious management measures. The present experiment was conducted on Citrus unshiu Marc. cv. ‘Miyagawa Wase’ over two successive seasons—2022 and 2023. The dynamic changes in fruit morphology were recorded using calipers, and the peel strength was assessed via a Plus Texture Analyzer. Paraffin sectioning technology was used to observe the morphological structure of peel cells. At 10 days after full bloom (DAFB), the peel cells exhibited vigorous proliferation, and the fruit and peel thicknesses underwent rapid expansion. At 50–60 d after full bloom, the longitudinal and transverse diameters of the fruit exhibited a marked increase in the growth rate of the former over the latter. At 80 d after full bloom, both the peel thickness change and the fruit growth rate exhibited a marked deceleration, and the albedo layer cells began to show signs of perforation. The following two time points were preliminarily proposed as the key points for the control of citrus fruit cracking: key point one was 50–60 days after full bloom; and key point two was 80–90 days after full bloom. The nitrogen (N), phosphorus (P), and potassium (K) contents in the different orchards were measured via the semi-micro Kjeldahl nitrogen method, the molybdenum–antimony colorimetric method, and flame photometry, respectively. The determination of other mineral elements was conducted by means of inductively coupled plasma spectroscopy. Principal component analysis was employed to analyze the 21-parameter indices of mineral elements in soil and leaf samples from the three orchards with different levels of fruit cracking. The study found that high concentrations of leaf Fe, P, and soil Cu, as well as organic matter content, contributed negatively to the extent of fruit cracking. The impact of diverse control measures on the incidence of fruit cracking was subsequently observed, following the implementation of tree crown spray treatments. The application of 0.5% calcium superphosphate and 0.006% EDTA-Fe, in combination with 10 ppm GA₃ sprayed during two critical periods, significantly reduced fruit cracking and did not adversely affect the internal or external quality of the fruits. The study emphasises the necessity of customising management measures according to the developmental characteristics of citrus fruits, given the observed varietal and regional distinctions in susceptibility to cracking. These findings are pivotal for advancing research in the field of fruit cracking and promoting the healthy development of the industry. Full article

Cultivated meat, an advancement in cellular agriculture, holds promise in addressing environmental, ethical, and health challenges associated with traditional meat production. Utilizing tissue engineering principles, cultivated meat production employs biomaterials and technologies to create cell-based structures by introducing cells into a biocompatible scaffold, mimicking tissue organization. Among the cell sources used for producing muscle-like tissue for cultivated meats, primary adult stem cells like muscle satellite cells exhibit robust capabilities for proliferation and differentiation into myocytes, presenting a promising avenue for cultivated meat production. Evolutionarily optimized for growth in a 3D microenvironment, these cells benefit from the biochemical and biophysical cues provided by the extracellular matrix (ECM), regulating cell organization, interactions, and behavior. While plant protein-based scaffolds have been explored for their utilization for cultivated meat, they lack the biological cues for animal cells unless functionalized. Conversely, a decellularized bovine placental tissue ECM, processed from discarded birth tissue, achieves the biological functionalities of animal tissue ECM without harming animals. In this study, collagen and total ECM were prepared from decellularized bovine placental tissues. The collagen content was determined to be approximately 70% and 40% in isolated collagen and ECM, respectively. The resulting porous scaffolds, crosslinked through a dehydrothermal (DHT) crosslinking method without chemical crosslinking agents, supported the growth of bovine myoblasts. ECM scaffolds exhibited superior compatibility and stability compared to collagen scaffolds. In an attempt to make cultivate meat constructs, bovine myoblasts were cultured in steak-shaped ECM scaffolds for about 50 days. The resulting construct not only resembled muscle tissues but also displayed high cellularity with indications of myogenic differentiation. Furthermore, the meat constructs were cookable and able to sustain the grilling/frying. Our study is the first to utilize a unique bovine placentome-derived ECM scaffold to create a muscle tissue-like meat construct, demonstrating a promising and sustainable option for cultivated meat production. Full article

Nowadays, it is important to create the optimal technology for the absorption of flue gases with high CO₂ content. In this regard, the aim of the investigation is to study the five different microalgae strains (Chlorella vulgaris, Chlorella ellipsoidea, Elliptochoris subsphaerica, Gloeotila pulchra, and Arthrospira platensis) under the influence of flue gases. The cultivation of microalgae was carried out in the atmosphere of flue gases with a gas flow rate of approximately 1 L·min⁻¹ at high CO₂ concentrations (3, 6, or 8%—from lower to higher concentrations), under continuous (24 h·d⁻¹) illumination intensity of 200 µmol quanta·m⁻²·s⁻¹ and a constant temperature of 27 ± 1 °C. The duration of the experiments was 12 days. Chlorella vulgaris and Chlorella ellipsoidea demonstrated the highest biomass growth rate at CO₂ = 6% (0.79 and 0.74 g·L⁻¹·d⁻¹, respectively). The lowest growth rate (0.21 g·L⁻¹·d⁻¹) was achieved for Arthrospira platensis at CO₂ = 3 and 6%. There was no significant drop in pH in the entire series of experiments. The results of microscopy showed a lack or a minimal number of dead cells in the strains under selected conditions. The obtained results can be used for further development of CO₂ capture and storage technologies. Full article

Cell co-culture technology aims to study the communication mechanism between cells and to better reveal the interactions and regulatory mechanisms involved in processes such as cell growth, differentiation, apoptosis, and other cellular activities. This is achieved by simulating the complex organismic environment. Such studies are of great significance for understanding the physiological and pathological processes of multicellular organisms. As an emerging cell cultivation technology, 3D cell co-culture technology, based on microfluidic chips, can efficiently, rapidly, and accurately achieve cell co-culture. This is accomplished by leveraging the unique microchannel structures and flow characteristics of microfluidic chips. The technology can simulate the native microenvironment of cell growth, providing a new technical platform for studying intercellular communication. It has been widely used in the research of oncology, immunology, neuroscience, and other fields. In this review, we summarize and provide insights into the design of cell co-culture systems on microfluidic chips, the detection methods employed in co-culture systems, and the applications of these models. Full article

Bacterial ghosts (BGs) are hollow bacterial cell envelopes with intact cellular structures, presenting as promising candidates for various biotechnological and biomedical applications. However, the yield and productivity of BGs have encountered limitations, hindering their large-scale preparation and multi-faceted applications of BGs. Further optimization of BGs is needed for the commercial application of BG technology. In this study, we screened out the most effective lysis protein ID52-E-W4A among 13 mutants based on phage ID52 lysis protein E and optimized the liquid culture medium for preparing Escherichia coli Nissle 1917 (EcN). The results revealed a significantly higher lysis rate of ID52-E-W4A compared to that of ID52-E in the 2xYT medium. Furthermore, EcN BGs were cultivated in a fermenter, achieving an initial OD₆₀₀ as high as 6.0 after optimization, indicating enhanced BG production. Moreover, the yield of ID52-E-W4A-induced BGs reached 67.0%, contrasting with only a 3.1% yield from φX174-E-induced BGs. The extended applicability of the lysis protein ID52-E-W4A was demonstrated through the preparation of Salmonella pullorum ghosts and Salmonella choleraesuis ghosts. Knocking out the molecular chaperone gene slyD and dnaJ revealed that ID52-mediated BGs could still undergo lysis. Conversely, overexpression of integral membrane enzyme gene mraY resulted in the loss of lysis activity for ID52-E, suggesting that the lysis protein ID52-E may no longer rely on SlyD or DnaJ to function, with MraY potentially being the target of ID52-E. This study introduces a novel approach utilizing ID52-E-W4A for recombinant expression, accelerating the BG formation and thereby enhancing BG yield and productivity. Full article

Biomimetic scaffolds imitate native tissue and can take a multidimensional form. They are biocompatible and can influence cellular metabolism, making them attractive bioengineering platforms. The use of biomimetic scaffolds adds complexity to traditional cell cultivation methods. The most commonly used technique involves cultivating cells on a flat surface in a two-dimensional format due to its simplicity. A three-dimensional (3D) format can provide a microenvironment for surrounding cells. There are two main techniques for obtaining 3D structures based on the presence of scaffolding. Scaffold-free techniques consist of spheroid technologies. Meanwhile, scaffold techniques contain organoids and all constructs that use various types of scaffolds, ranging from decellularized extracellular matrix (dECM) through hydrogels that are one of the most extensively studied forms of potential scaffolds for 3D culture up to 4D bioprinted biomaterials. 3D bioprinting is one of the most important techniques used to create biomimetic scaffolds. The versatility of this technique allows the use of many different types of inks, mainly hydrogels, as well as cells and inorganic substances. Increasing amounts of data provide evidence of vast potential of biomimetic scaffolds usage in tissue engineering and personalized medicine, with the main area of potential application being the regeneration of skin and musculoskeletal systems. Recent papers also indicate increasing amounts of in vivo tests of products based on biomimetic scaffolds, which further strengthen the importance of this branch of tissue engineering and emphasize the need for extensive research to provide safe for humansbiomimetic tissues and organs. In this review article, we provide a review of the recent advancements in the field of biomimetic scaffolds preceded by an overview of cell culture technologies that led to the development of biomimetic scaffold techniques as the most complex type of cell culture. Full article

Shape-controlled 3D tissues resemble natural living tissues in human and animal bodies and are essential materials for developing and improving technologies in regenerative medicine, drug discovery, and biological robotics. In previous studies, shape-controlled 3D tissues were fabricated using scaffold structures or 3D bioprinting techniques. However, controlling the shape of 3D tissues without leaving non-natural materials inside the 3D tissue and efficiently fabricating them remains challenging. In this paper, we propose a novel method for fabricating shape-controlled 3D tissues free of non-natural materials using a flexible high-porosity porous structure (HPPS). The HPPS consisted of a micromesh with pore sizes of 14.87 ± 1.83 μm, lattice widths of 2.24 ± 0.10 μm, thicknesses of 9.96 ± 0.92 μm, porosity of 69.06 ± 3.30%, and an I-shaped microchamber of depth 555.26 ± 11.17 μm. U-87 human glioma cells were cultured in an I-shaped HPPS microchamber for 48 h. After cultivation, the 3D tissue was released within a few seconds while maintaining its I-shape. Specific chemicals, such as proteolytic enzymes, were not used. Moreover, the viability of the released cells composed of shape-controlled 3D tissues free of non-natural materials was above 90%. Therefore, the proposed fabrication method is recommended for shape-controlled 3D tissues free of non-natural materials without applying significant stresses to the cells. Full article

Bioprinting provides a powerful tool for regenerative medicine, as it allows tissue construction with a patient’s specific geometry. However, tissue culture and maturation, commonly supported by dynamic bioreactors, are needed. We designed a workflow that creates an implant-specific bioreactor system, which is easily producible and customizable and supports cell cultivation and tissue maturation. First, a bioreactor was designed and different tissue geometries were simulated regarding shear stress and nutrient distribution to match cell culture requirements. These tissues were then directly bioprinted into the 3D-printed bioreactor. To prove the ability of cell maintenance, C2C12 cells in two bioinks were printed into the system and successfully cultured for two weeks. Next, human mesenchymal stem cells (hMSCs) were successfully differentiated toward an adipocyte lineage. As the last step of the presented strategy, we developed a prototype of an automated mobile docking station for the bioreactor. Overall, we present an open-source bioreactor system that is adaptable to a wound-specific geometry and allows cell culture and differentiation. This interdisciplinary roadmap is intended to close the gap between the lab and clinic and to integrate novel 3D-printing technologies for regenerative medicine. Full article

Cultivated meat is a nascent technology that aims to create an environmentally and animal-friendly alternative to conventional meat. Producing skeletal muscle tissue in an animal-free system allowing for high levels of myofusion and maturation is important for the nutritional and sensorial value of cultivated meat. Alginate is an attractive biomaterial to support muscle formation as it is food-safe, sustainable and cheap and can be crosslinked using non-toxic methods. Although alginate can be functionalized to promote cell attachment, limitations in its mechanical properties, including form, viscosity, and stress relaxation, hinder the cellular capacity for myogenic differentiation and maturation in alginate-based hydrogels. Here, we show that the addition of electrospun short-stranded zein fibers increased hydrogel degradation, resulting in faster compaction, improved cell–gel interaction, and enhanced alignment of bovine muscle precursor cells. We conclude that fiber-hydrogel composites are a promising approach to support optimal formation of 3D constructs, by improving tissue stability and thus prolonging culture duration. Together, this improves muscle-related protein content by facilitating myogenic differentiation and priming muscle organoids for maturation. Full article

In modern energy, various technologies for absorbing carbon dioxide from the atmosphere are being considered, including photosynthetic microalgae. An important task is to obtain maximum productivity at high concentrations of CO₂ in gas–air mixtures. In this regard, the aim of the investigation is to study the effect of light intensity on the biomass growth and biochemical composition of five different microalgae strains: Arthrospira platensis, Chlorella ellipsoidea, Chlorella vulgaris, Gloeotila pulchra, and Elliptochloris subsphaerica. To assess the viability of microalgae cells, the method of cytochemical staining with methylene blue, which enables identifying dead cells during microscopy, was used. The microalgae were cultivated at 6% CO₂ and five different intensities: 80, 120, 160, 200, and 245 μmol quanta·m⁻²·s⁻¹. The maximum growth rate among all strains was obtained for C. vulgaris (0.78 g·L⁻¹·d⁻¹) at an illumination intensity of 245 µmol quanta·m⁻²·s⁻¹. For E. subsphaerica and A. platensis, similar results (approximately 0.59 and 0.25 g·L⁻¹·d⁻¹ for each strain) were obtained at an illumination intensity of 160 and 245 µmol quanta·m⁻²·s⁻¹. A decrease in protein content with an increase in illumination was noted for C. vulgaris (from 61.0 to 46.6%) and A. platensis (from 43.8 to 33.6%), and a slight increase in lipid content was shown by A. platensis (from 17.8 to 21.4%). The possibility of increasing microalgae biomass productivity by increasing illumination has been demonstrated. This result can also be considered as showing potential for enhanced lipid microalgae production for biodiesel applications. Full article