bioreactor culture
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2021 ◽  
Vol 22 (24) ◽  
pp. 13549
Author(s):  
Sadman Sakib ◽  
Anna Voigt ◽  
Nathalia de Lima e Martins Lara ◽  
Lin Su ◽  
Mark Ungrin ◽  
...  

Male survivors of childhood cancer are at risk of suffering from infertility in adulthood because of gonadotoxic chemotherapies. For adult men, sperm collection and preservation are routine procedures prior to treatment; however, this is not an option for pre-pubertal children. From young boys, a small biopsy may be taken before chemotherapy, and spermatogonia may be propagated in vitro for future transplantation to restore fertility. A robust system that allows for scalable expansion of spermatogonia within a controlled environment is therefore required. Stirred suspension culture has been applied to different types of stem cells but has so far not been explored for spermatogonia. Here, we report that pre-pubertal porcine spermatogonia proliferate more in bioreactor suspension culture, compared with static culture. Interestingly, oxygen tension provides an avenue to modulate spermatogonia status, with culture under 10% oxygen retaining a more undifferentiated state and reducing proliferation in comparison with the conventional approach of culturing under ambient oxygen levels. Spermatogonia grown in bioreactors upregulate the Wnt/ β-catenin pathway, which, along with enhanced gas and nutrient exchange observed in bioreactor culture, may synergistically account for higher spermatogonia proliferation. Therefore, stirred suspension bioreactors provide novel platforms to culture spermatogonia in a scalable manner and with minimal handling.


2021 ◽  
Vol 11 (24) ◽  
pp. 11819
Author(s):  
Katarzyna Wierzchowska ◽  
Bartłomiej Zieniuk ◽  
Dorota Nowak ◽  
Agata Fabiszewska

Microbial lipids called a sustainable alternative to traditional vegetable oils invariably capture the attention of researchers. In this study, the effect of limiting inorganic phosphorus (KH2PO4) and nitrogen ((NH4)2SO4) sources in lipid-rich culture medium on the efficiency of cellular lipid biosynthesis by Y. lipolytica yeast has been investigated. In batch cultures, the carbon source was rapeseed waste post-frying oil (50 g/dm3). A significant relationship between the concentration of KH2PO4 and the amount of lipids accumulated has been revealed. In the shake-flask cultures, storage lipid yield was correlated with lower doses of phosphorus source in the medium. In bioreactor culture in mineral medium with (g/dm3) 3.0 KH2PO4 and 3.0 (NH4)2SO4, the cellular lipid yield was 47.5% (w/w). Simultaneous limitation of both phosphorus and nitrogen sources promoted lipid accumulation in cells, but at the same time created unfavorable conditions for biomass growth (0.78 gd.m./dm3). Increased phosphorus availability with limited cellular access to nitrogen resulted in higher biomass yields (7.45 gd.m./dm3) than phosphorus limitation in a nitrogen-rich medium (4.56 gd.m./dm3), with comparable lipid yields (30% and 32%). Regardless of the medium composition, the yeast preferentially accumulated oleic and linoleic acids as well as linolenic acid up to 8.89%. Further, it is crucial to determine the correlation between N/P molar ratios, biomass growth and efficient lipid accumulation. In particular, considering the contribution of phosphorus as a component of coenzymes in many metabolic pathways, including lipid biosynthesis and respiration processes, its importance as a factor in the cultivation of the oleaginous microorganisms was highlighted.


Agronomy ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 2414
Author(s):  
Anna De De Carlo ◽  
Waed Tarraf ◽  
Maurizio Lambardi ◽  
Carla Benelli

The cultivation of medicinal plants and the production of bioactive compounds derived from them are of fundamental importance and interest, not only at the pharmacological level but also in nutraceutical and cosmetic industries and in functional foods, as well as plant protection in agriculture. In order to respond adequately to the increased demands of the global market from a quantitative and qualitative point of view and to guarantee environmental sustainability of the productions, it is necessary to resort to innovation tools, such as tissue culture in vitro technology. Nowadays, it is well known that the cultivation through the Temporary Immersion System (TIS) in a bioreactor has considerable advantages both for the in vitro mass production of the plants and for the production of secondary metabolites. The present review focuses on the application of TIS during the last two decades to produce biomass and bioactive compounds from medicinal plants. Indeed, almost one hundred papers are discussed, and they particularly focus on the effects of the culture system, vessel design and equipment, immersion time and frequency, and substrate composition for 88 medicinal species in TIS bioreactor culture.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rupak Dua ◽  
Hugh Jones ◽  
Philip C. Noble

AbstractRecent advances in materials and manufacturing processes have allowed the fabrication of intricate implant surfaces to facilitate bony attachment. However, refinement and evaluation of these new design strategies are hindered by the cost and complications of animal studies, particularly during early iterations in the development process. To address this problem, we have previously constructed and validated an ex-vivo bone bioreactor culture system that can maintain the viability of bone samples for an extended period ex-vivo. In this study, we investigated the mineralization of a titanium wire mesh scaffold under both static and dynamic culturing using our ex vivo bioreactor system. Thirty-six cancellous bone cores were harvested from bovine metatarsals at the time of slaughter and divided into five groups under the following conditions: Group 1) Isolated bone cores placed in static culture, Group 2) Unloaded bone cores placed in static culture in contact with a fiber-mesh metallic scaffold, Group 3) Bone cores placed in contact with a fiber-mesh metallic scaffold under the constant pressure of 150 kPa, Group 4) Bone core placed in contact with a fiber-mesh metallic scaffold and exposed to cyclic loading with continuous perfusion flow of media within the ex-vivo culture system and Group 5) Bone core evaluated on Day 0 to serve as a positive control for comparison with all other groups at weeks 4 and 7. Bone samples within Groups 1–4 were incubated for 4 and 7 weeks and then evaluated using histological examination (H&E) and the Live-Dead assay (Life Technologies). Matrix deposits on the metallic scaffolds were examined with scanning electron microscopy (SEM), while the chemical composition of the matrix was measured using energy-dispersive x-ray spectroscopy (EDX). We found that the viability of bone cores was maintained after seven weeks of loading in our ex vivo system. In addition, SEM images revealed crystallite-like structures on the dynamically loaded metal coupons (Group 4), corresponding to the initial stages of mineralization. EDX results further confirmed the presence of carbon at the interface and calcium phosphates in the matrix. We conclude that a bone bioreactor can be used as an alternate tool for in-vivo bone ingrowth studies of new implant surfaces or coatings.


2021 ◽  
pp. 002203452110288
Author(s):  
W. Ma ◽  
H. Lyu ◽  
M. Pandya ◽  
G. Gopinathan ◽  
X. Luan ◽  
...  

The nervous system exerts finely tuned control over all aspects of the life of an organism, including pain, sensation, growth, and development. Recent developments in tissue regeneration research have increasingly turned to small molecule peptides to tailor and augment the biological response following tissue loss or injury. In the present study, we have introduced the small molecule peptide galanin (GAL) as a novel scaffold-coating agent for the healing and regeneration of craniofacial tissues. Using immunohistochemistry, we detected GAL and GAL receptors in healthy periodontal tissues and in the proximity of blood vessels, while exposure to our periodontal disease regimen resulted in a downregulation of GAL. In a 3-dimensional bioreactor culture, GAL coating of collagen scaffolds promoted cell proliferation and matrix synthesis. Following subcutaneous implantation, GAL-coated scaffolds were associated with mineralized bone-like tissue deposits, which reacted positively for alizarin red and von Kossa, and demonstrated increased expression and protein levels of RUNX2, OCN, OSX, and iBSP. In contrast, the GAL receptor antagonist galantide blocked the effect of GAL on Runx2 expression and inhibited mineralization in our subcutaneous implantation model. Moreover, GAL coating promoted periodontal regeneration and a rescue of the periodontal defect generated in our periodontitis model mice. Together, these data demonstrate the efficacy of the neuropeptide GAL as a coating material for tissue regeneration. They are also suggestive of a novel role for neurogenic signaling pathways in craniofacial and periodontal regeneration.


Author(s):  
Hamza Patel ◽  
Peter Archibald ◽  
Cindy Jung ◽  
Blair Roszell ◽  
Farlan Veraitch ◽  
...  

Lentiviral vectors (LVV) represent an important tool for vaccine development and other therapeutic modalities. However, inefficiencies in LVV manufacturing processes, such as the inability to achieve high cell densities with HEK293T cell lines in a fed batch process, have resulted in poor upstream yields. Optimisation of cell culture conditions is needed to improve upstream yields, which can be expedited by high-throughput screening (HTP). In this work, we describe the use of the 24 deep square well (24-DSW) microwell platform to develop a scale-down mimic of GSK’s established stable suspension LVV production process model at 2 L bioreactor scale. We found that matched mixing time was an effective basis for scale-translation between the stirred tank reactor (STR) and microwells. The growth kinetics and LVV productivity profile in the microwell were reproducible and comparable to the 2 L bioreactor process model. In both vessels, a 6-fold increase in cell density was achieved at the harvest time point and high cell viability (i.e. > 90 %) was also maintained throughout the entirety of the cultures. The 24-DSW model, therefore, is an effective scale-down model for larger-scale stirred-tank bioreactor culture and provides an important tool for rapid, high-throughput optimization of the LVV production process.


2021 ◽  
Vol 22 (15) ◽  
pp. 7920
Author(s):  
Myroslava Mytsyk ◽  
Giulia Cerino ◽  
Gregory Reid ◽  
Laia Gili Sole ◽  
Friedrich S. Eckstein ◽  
...  

The therapeutic potential of mesenchymal stromal/stem cells (MSC) for treating cardiac ischemia strongly depends on their paracrine-mediated effects and their engraftment capacity in a hostile environment such as the infarcted myocardium. Adipose tissue-derived stromal vascular fraction (SVF) cells are a mixed population composed mainly of MSC and vascular cells, well known for their high angiogenic potential. A previous study showed that the angiogenic potential of SVF cells was further increased following their in vitro organization in an engineered tissue (patch) after perfusion-based bioreactor culture. This study aimed to investigate the possible changes in the cellular SVF composition, in vivo angiogenic potential, as well as engraftment capability upon in vitro culture in harsh hypoxia conditions. This mimics the possible delayed vascularization of the patch upon implantation in a low perfused myocardium. To this purpose, human SVF cells were seeded on a collagen sponge, cultured for 5 days in a perfusion-based bioreactor under normoxia or hypoxia (21% and <1% of oxygen tension, respectively) and subcutaneously implanted in nude rats for 3 and 28 days. Compared to ambient condition culture, hypoxic tension did not alter the SVF composition in vitro, showing similar numbers of MSC as well as endothelial and mural cells. Nevertheless, in vitro hypoxic culture significantly increased the release of vascular endothelial growth factor (p < 0.001) and the number of proliferating cells (p < 0.00001). Moreover, compared to ambient oxygen culture, exposure to hypoxia significantly enhanced the vessel length density in the engineered tissues following 28 days of implantation. The number of human cells and human proliferating cells in hypoxia-cultured constructs was also significantly increased after 3 and 28 days in vivo, compared to normoxia. These findings show that a possible in vivo delay in oxygen supply might not impair the vascularization potential of SVF- patches, which qualifies them for evaluation in a myocardial ischemia model.


Antibiotics ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 886
Author(s):  
Youngbeom Ahn ◽  
Ji Young Jung ◽  
Ohgew Kweon ◽  
Brian T. Veach ◽  
Sangeeta Khare ◽  
...  

Studying potential dietary exposure to antimicrobial drug residues via meat and dairy products is essential to ensure human health and consumer safety. When studying how antimicrobial residues in food impact the development of antimicrobial drug resistance and disrupt normal bacteria community structure in the intestine, there are diverse methodological challenges to overcome. In this study, traditional cultures and molecular analysis techniques were used to determine the effects of tetracycline at chronic subinhibitory exposure levels on human intestinal microbiota using an in vitro continuous flow bioreactor. Six bioreactor culture vessels containing human fecal suspensions were maintained at 37 °C for 7 days. After a steady state was achieved, the suspensions were dosed with 0, 0.015, 0.15, 1.5, 15, or 150 µg/mL tetracycline, respectively. Exposure to 150 µg/mL tetracycline resulted in a decrease of total anaerobic bacteria from 1.9 × 107 ± 0.3 × 107 down to 2 × 106 ± 0.8 × 106 CFU/mL. Dose-dependent effects of tetracycline were noted for perturbations of tetB and tetD gene expression and changes in acetate and propionate concentrations. Although no-observed-adverse-effect concentrations differed, depending on the traditional cultures and the molecular analysis techniques used, this in vitro continuous flow bioreactor study contributes to the knowledge base regarding the impact of chronic exposure of tetracycline on human intestinal microbiota.


2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Maki Moriwaki-Takano ◽  
Chikako Asada ◽  
Yoshitosi Nakamura

AbstractSpiculisporic acid (SA) is a fatty acid-type biosurfactant with one lactone ring and two carboxyl groups. It has been used in metal removers and cosmetics, because of its low propensity to cause irritation to the skin, its anti-bacterial properties, and high surface activity. In the present study, we report an effective method for producing SA by selecting a high-producing strain and investigating the effective medium components, conditions, and environments for its culture. Among the 11 kinds of Talaromyces species, T. trachyspermus NBRC 32238 showed the highest production of a crystalline substance, which was determined to be SA using NMR. The strain was able to produce SA under acidic conditions from hexoses, pentoses, and disaccharides, with glucose and sucrose serving as the most appropriate substrates. Investigation of nitrogen sources and trace metal ions revealed meat extract and FeCl3 as components that promoted SA production. Upon comparing the two types of cultures with glucose in a baffle flask or aeration bioreactor, SA production was found to be slightly higher in the flask than in the reactor. In the bioreactor culture, sucrose was found to be an appropriate substrate for SA production, as compared to glucose, because with sucrose, the lag time until the start of SA production was shortened. Finally, fed-batch culture with sucrose resulted in 60 g/L of SA, with a total yield of 0.22 g SA/g sucrose and a productivity of 6.6 g/L/day.


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