scholarly journals Identification, Selection, and Enrichment of Cardiomyocyte Precursors

2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
Bianca Ferrarini Zanetti ◽  
Walter José Gomes ◽  
Sang Won Han
Keyword(s):  
Tissue Engineering ◽  
Cell Therapy ◽  
Large Scale ◽  
Cellular Reprogramming ◽  
Efficient Production ◽  
Scale Production ◽  
Protein Markers ◽  
Large Numbers ◽  
Large Scale Production ◽  
Cell Doubling Time

The large-scale production of cardiomyocytes is a key step in the development of cell therapy and tissue engineering to treat cardiovascular diseases, particularly those caused by ischemia. The main objective of this study was to establish a procedure for the efficient production of cardiomyocytes by reprogramming mesenchymal stem cells from adipose tissue. First, lentiviral vectors expressing neoR and GFP under the control of promoters expressed specifically during cardiomyogenesis were constructed to monitor cell reprogramming into precardiomyocytes and to select cells for amplification and characterization. Cellular reprogramming was performed using 5′-azacytidine followed by electroporation with plasmid pOKS2a, which expressed Oct4, Sox2, and Klf4. Under these conditions, GFP expression began only after transfection with pOKS2a, and less than 0.015% of cells were GFP+. These GFP+cells were selected for G418 resistance to find molecular markers of cardiomyocytes by RT-PCR and immunocytochemistry. Both genetic and protein markers of cardiomyocytes were present in the selected cells, with some variations among them. Cell doubling time did not change after selection. Together, these results indicate that enrichment with vectors expressing GFP and neoR under cardiomyocyte-specific promoters can produce large numbers of cardiomyocyte precursors (CMPs), which can then be differentiated terminally for cell therapy and tissue engineering.

scholarly journals Efficient Production of l-Ribose with a Recombinant Escherichia coli Biocatalyst

2008 ◽  
Vol 74 (10) ◽  
pp. 2967-2975 ◽  
Author(s):  
Ryan D. Woodyer ◽  
Nathan J. Wymer ◽  
F. Michael Racine ◽  
Shama N. Khan ◽  
Badal C. Saha
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Active Form ◽  
Apium Graveolens ◽  
Efficient Production ◽  
Scale Production ◽  
Gene Encoding ◽  
Large Scale Production ◽  
One Step ◽  
Rare Sugars

ABSTRACT A new synthetic platform with potential for the production of several rare sugars, with l-ribose as the model target, is described. The gene encoding the unique NAD-dependent mannitol-1-dehydrogenase (MDH) from Apium graveolens (garden celery) was synthetically constructed for optimal expression in Escherichia coli. This MDH enzyme catalyzes the interconversion of several polyols and their l-sugar counterparts, including the conversion of ribitol to l-ribose. Expression of recombinant MDH in the active form was successfully achieved, and one-step purification was demonstrated. Using the created recombinant E. coli strain as a whole-cell catalyst, the synthetic utility was demonstrated for production of l-ribose, and the system was improved using shaken flask experiments. It was determined that addition of 50 to 500 μM ZnCl2 and addition of 5 g/liter glycerol both improved production. The final levels of conversion achieved were >70% at a concentration of 40 g/liter and >50% at a concentration of 100 g/liter. The best conditions determined were then scaled up to a 1-liter fermentation that resulted in 55% conversion of 100 g/liter ribitol in 72 h, for a volumetric productivity of 17.4 g liter−1 day−1. This system represents a significantly improved method for the large-scale production of l-ribose.

Vegetative propagation of Sitka spruce

1987 ◽  
Vol 93 (1-2) ◽  
pp. 197-203
Author(s):  
Allan John ◽  
Bill Mason
Keyword(s):  
Tissue Culture ◽  
Large Scale ◽  
Sitka Spruce ◽  
Scale Production ◽  
Stem Cuttings ◽  
Culture Techniques ◽  
Commercial Trial ◽  
Large Numbers ◽  
Large Scale Production ◽  
Tissue Culture Techniques

SynopsisA combination of two vegetative techniques is seen as a possibility for large-scale production of juvenile, rooted Sitka spruce cuttings of improved genotype. Tissue culture techniques, under development, would be used to produce large numbers of stock plants for stem cuttings production. Cuttings techniques, currently under commercial trial, would be used to produce the rooted plants for forest establishment.

scholarly journals Automated Large-Scale Production of Paclitaxel Loaded Mesenchymal Stromal Cells for Cell Therapy Applications

Pharmaceutics ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 411 ◽  
Author(s):  
Daniela Lisini ◽  
Sara Nava ◽  
Simona Frigerio ◽  
Simona Pogliani ◽  
Guido Maronati ◽  
...  
Keyword(s):  
Cell Therapy ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Large Scale ◽  
Good Manufacturing Practice ◽  
Scale Production ◽  
Large Scale Production ◽  
Advanced Therapies ◽  
Oncological Diseases

Mesenchymal stromal cells (MSCs) prepared as advanced therapies medicinal products (ATMPs) have been widely used for the treatment of different diseases. The latest developments concern the possibility to use MSCs as carrier of molecules, including chemotherapeutic drugs. Taking advantage of their intrinsic homing feature, MSCs may improve drugs localization in the disease area. However, for cell therapy applications, a significant number of MSCs loaded with the drug is required. We here investigate the possibility to produce a large amount of Good Manufacturing Practice (GMP)-compliant MSCs loaded with the chemotherapeutic drug Paclitaxel (MSCs-PTX), using a closed bioreactor system. Cells were obtained starting from 13 adipose tissue lipoaspirates. All samples were characterized in terms of number/viability, morphology, growth kinetics, and immunophenotype. The ability of MSCs to internalize PTX as well as the antiproliferative activity of the MSCs-PTX in vitro was also assessed. The results demonstrate that our approach allows a large scale expansion of cells within a week; the MSCs-PTX, despite a different morphology from MSCs, displayed the typical features of MSCs in terms of viability, adhesion capacity, and phenotype. In addition, MSCs showed the ability to internalize PTX and finally to kill cancer cells, inhibiting the proliferation of tumor lines in vitro. In summary our results demonstrate for the first time that it is possible to obtain, in a short time, large amounts of MSCs loaded with PTX to be used in clinical trials for the treatment of patients with oncological diseases.

Secondary Industries for Tropical Africa

Africa ◽  
1943 ◽  
Vol 14 (4) ◽  
pp. 170-176 ◽  
Author(s):  
Theresa Cahan
Keyword(s):  
Large Scale ◽  
Good Effect ◽  
Scale Production ◽  
Tropical Africa ◽  
Large Numbers ◽  
Large Scale Production ◽  
Initial Stage ◽  
The One ◽  
Opening Paragraph ◽  
Laborious Method

Opening ParagraphThere is a tendency in some quarters to regard secondary industries as a panacea for all the economic ills of tropical Africa. It would be well at this initial stage to sound a note of warning. In the past, the industrialization of agricultural countries has had two results: one good, one bad. On the one hand, the establishment in a country of labour-saving machinery and large-scale production in place of the old laborious method of making things by hand has led to a rise in the general standard of living within the country in terms of real incomes. On the other hand, the drift of workers to the towns and the herding together of large numbers of people in factories resulted in the sum of social evils associated with the ‘dark satanic mills’: overcrowding, sweated labour, destitution, unemployment, and many more. The problem for tropical Africa to-day is to combine the maximum of the first and good effect with a minimum of the second evil.

scholarly journals Replacement of a Metabolic Pathway for Large-Scale Production of Lactic Acid from Engineered Yeasts

1999 ◽  
Vol 65 (9) ◽  
pp. 4211-4215 ◽  
Author(s):  
Danilo Porro ◽  
Michele M. Bianchi ◽  
Luca Brambilla ◽  
Rossella Menghini ◽  
Davide Bolzani ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Large Scale ◽  
Kluyveromyces Lactis ◽  
Lactic Acid Production ◽  
Pyruvate Decarboxylase ◽  
Polymer Materials ◽  
Efficient Production ◽  
Scale Production ◽  
High Productivity ◽  
Large Scale Production

ABSTRACT Interest in the production of l-(+)-lactic acid is presently growing in relation to its applications in the synthesis of biodegradable polymer materials. With the aim of obtaining efficient production and high productivity, we introduced the bovinel-lactate dehydrogenase gene (LDH) into a wild-type Kluyveromyces lactis yeast strain. The observed lactic acid production was not satisfactory due to the continued coproduction of ethanol. A further restructuring of the cellular metabolism was obtained by introducing the LDH gene into aK. lactis strain in which the unique pyruvate decarboxylase gene had been deleted. With this modified strain, in which lactic fermentation substituted completely for the pathway leading to the production of ethanol, we obtained concentrations, productivities, and yields of lactic acid as high as 109 g liter−1, 0.91 g liter−1 h−1, and 1.19 mol per mole of glucose consumed, respectively. The organic acid was also produced at pH levels lower than those usual for bacterial processes.

scholarly journals Enhancing Stem Cell Therapy for Cartilage Repair in Osteoarthritis—A Hydrogel Focused Approach

Gels ◽  
2021 ◽  
Vol 7 (4) ◽  
pp. 263
Author(s):  
Yisi Liu ◽  
Meng Wang ◽  
Yixuan Luo ◽  
Qianyi Liang ◽  
Yin Yu ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Therapy ◽  
Cell Fate ◽  
Cartilage Repair ◽  
Stem Cell Therapy ◽  
Large Scale ◽  
Scale Production ◽  
Cell Based Therapy ◽  
Large Scale Production ◽  
Tissue Site

Sem cells hold tremendous promise for the treatment of cartilage repair in osteoarthritis. In addition to their multipotency, stem cells possess immunomodulatory effects that can alleviate inflammation and enhance cartilage repair. However, the widely clinical application of stem cell therapy to cartilage repair and osteoarthritis has proven difficult due to challenges in large-scale production, viability maintenance in pathological tissue site and limited therapeutic biological activity. This review aims to provide a perspective from hydrogel-focused approach to address few key challenges in stem cell-based therapy for cartilage repair and highlight recent progress in advanced hydrogels, particularly microgels and dynamic hydrogels systems for improving stem cell survival, retention and regulation of stem cell fate. Finally, progress in hydrogel-assisted gene delivery and genome editing approaches for the development of next generation of stem cell therapy for cartilage repair in osteoarthritis are highlighted.

scholarly journals Morphology, mechanical strength and degradation of polyhydroxyalkanoate scaffolds

2018 ◽  
Vol 27 (48) ◽  
Author(s):  
Liliana Maria Arroyave-Muñoz ◽  
Claudia Patricia Ossa-Orozco
Keyword(s):  
Tissue Engineering ◽  
Articular Cartilage ◽  
Mechanical Strength ◽  
Large Scale ◽  
Freezing Process ◽  
Scale Production ◽  
Natural Polymers ◽  
Large Scale Production ◽  
Cartilage Injuries ◽  
Innovative Material

Tissue engineering (TE) seeks to improve the unsatisfactory development of implants and medical procedures to solve bone and cartilage injuries. TE aims at regenerating tissues using cell growth platforms (scaffolds), which may consist of natural polymers such as polyhydroxyalkanoate (PHA). PHA is an innovative material useful in medical applications due to its degradation capability and bacterial origin that allows large-scale production and control final properties. In this research, we developed commercial PHA scaffolds using the lyophilization technique with a factorial experimental design. We used dichloromethane as PHA solvent, tergitol as surfactant, and liquid nitrogen (N2) for the freezing process. We characterized the PHA by Fourier-transform infrared spectroscopy (FTIR) and thermogravimetric analysis (TGA); and the scaffolds by scanning electron microscopy (SEM) and mechanical compression and hydrolysis degradation tests. The characterization of the PHA indicated that the material is a mixture of PHA and polylactic acid (PLA). The results showed a suitable pore distribution for migration of chondrocytes through the scaffold, in addition to a behavior similar to that of the articular cartilage, although it presented lower mechanical strength. Also, the scaffolds displayed mass loss in a non-linear way related to the percentage of PHA present in the sample. In conclusion, PHA scaffolds have a potential use in tissue engineering for restoring articular cartilage.

scholarly journals Tolerance engineering in Deinococcus geothermalis by heterologous efflux pumps

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Erika Boulant ◽  
Emmanuelle Cambon ◽  
Julia Vergalli ◽  
Rémi Bernard ◽  
Fabienne Neulat-Ripoll ◽  
...  
Keyword(s):  
Large Scale ◽  
Efflux Pumps ◽  
Toxic Effects ◽  
Efficient Production ◽  
Scale Production ◽  
Gene Encoding ◽  
Large Scale Production ◽  
Deinococcus Geothermalis ◽  
Genes Encoding ◽  
Susceptibility Tests

AbstractProducing industrially significant compounds with more environmentally friendly represents a challenging task. The large-scale production of an exogenous molecule in a host microfactory can quickly cause toxic effects, forcing the cell to inhibit production to survive. The key point to counter these toxic effects is to promote a gain of tolerance in the host, for instance, by inducing a constant flux of the neo-synthetized compound out of the producing cells. Efflux pumps are membrane proteins that constitute the most powerful mechanism to release molecules out of cells. We propose here a new biological model, Deinococcus geothermalis, organism known for its ability to survive hostile environment; with the aim of coupling the promising industrial potential of this species with that of heterologous efflux pumps to promote engineering tolerance. In this study, clones of D. geothermalis containing various genes encoding chromosomal heterologous efflux pumps were generated. Resistant recombinants were selected using antibiotic susceptibility tests to screen promising candidates. We then developed a method to determine the efflux efficiency of the best candidate, which contains the gene encoding the MdfA of Salmonella enterica serovar Choleraesuis. We observe 1.6 times more compound in the external medium of the hit recombinant than that of the WT at early incubation time. The data presented here will contribute to better understanding of the parameters required for efficient production in D. geothermalis.

scholarly journals Production of 10-methyl branched fatty acids in yeast

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Hannah G. Blitzblau ◽  
Andrew L. Consiglio ◽  
Paulo Teixeira ◽  
Donald V. Crabtree ◽  
Shuyan Chen ◽  
...  
Keyword(s):  
Fatty Acids ◽  
High Performance ◽  
Large Scale ◽  
Unsaturated Fatty Acids ◽  
Acyl Chain ◽  
Efficient Production ◽  
Scale Production ◽  
Branched Fatty Acids ◽  
Large Scale Production ◽  
Methyl Branched Fatty Acids

Abstract Background Despite the environmental value of biobased lubricants, they account for less than 2% of global lubricant use due to poor thermo-oxidative stability arising from the presence of unsaturated double bonds. Methyl branched fatty acids (BFAs), particularly those with branching near the acyl-chain mid-point, are a high-performance alternative to existing vegetable oils because of their low melting temperature and full saturation. Results We cloned and characterized two pathways to produce 10-methyl BFAs isolated from actinomycetes and γ-proteobacteria. In the two-step bfa pathway of actinomycetes, BfaB methylates Δ9 unsaturated fatty acids to form 10-methylene BFAs, and subsequently, BfaA reduces the double bond to produce a fully saturated 10-methyl branched fatty acid. A BfaA-B fusion enzyme increased the conversion efficiency of 10-methyl BFAs. The ten-methyl palmitate production (tmp) pathway of γ-proteobacteria produces a 10-methylene intermediate, but the TmpA putative reductase was not active in E. coli or yeast. Comparison of BfaB and TmpB activities revealed a range of substrate specificities from C14-C20 fatty acids unsaturated at the Δ9, Δ10 or Δ11 position. We demonstrated efficient production of 10-methylene and 10-methyl BFAs in S. cerevisiae by secretion of free fatty acids and in Y. lipolytica as triacylglycerides, which accumulated to levels more than 35% of total cellular fatty acids. Conclusions We report here the characterization of a set of enzymes that can produce position-specific methylene and methyl branched fatty acids. Yeast expression of bfa enzymes can provide a platform for the large-scale production of branched fatty acids suitable for industrial and consumer applications.

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