scholarly journals Optimization of environment for high density Vero cell culture: effect of dissolved oxygen and nutrient supply on cell growth and changes in metabolites

1986 ◽  
Vol 81 (1) ◽  
pp. 65-103
Author(s):  
A.T. Nahapetian ◽  
J.N. Thomas ◽  
W.G. Thilly
Keyword(s):  
Cell Growth ◽  
Dissolved Oxygen ◽  
Vero Cell ◽  
Cell Density ◽  
Lactate Production ◽  
Nutrient Supply ◽  
High Density ◽  
Limiting Factors ◽  
Utilization Rate ◽  
Maximum Cell Density

This study was initiated for optimization of the environment of a technologically useful mammalian cell line for high density production. Cultures of Vero cells on microcarriers were perfused with 100%, 50%, 25% and 12.5% modified L15 media (galactose was replaced with 10 mM-fructose, with 4 mM-glutamine and 5% foetal bovine serum) in phosphate-buffered saline at either 4 or 8 vol. day-1. Cell growth, pH, dissolved oxygen, and changes in the metabolites, lactate to pyruvate and lactate to ammonia indices, demonstrated that under the conditions used in the present study, perfusion of cultures with 50% L15 medium in PBS at 8 vol. day-1 provided the optimum microenvironment for Vero cell growth. The highest cell density in the perfused cultures was 3 X 10(7) cells ml-1, which at these conditions was ten times higher than the maximum cell density (3 X 10(6) cells ml-1) obtained in a batch culture. Nutrient supply and conditioning factors were the most probable growth-limiting factors in cultures that were perfused with 12.5% and 25% L15 media, while multilayering, limitation of available oxygen, and accumulation of metabolic end products in the cellular microenvironment were the most probable causes of a density-dependent inhibition of cell growth observed under the optimized and overfed (supply of 100% L15 medium at the rate of 8 vol. day-1) culture conditions. Under the optimized environmental condition, the major source of energy was probably glutamine during the first week. However, significant utilization of fructose became evident at higher cell densities during the second week, when lactate production dramatically declined and reached an almost undetectable level, while respiration progressively assumed the predominant role in energy production. It is postulated that ‘available’ oxygen in the multicell-layered microenvironment of the optimized cultures was higher than in the overfed culture due to the greater utilization rate of oxygen for oxidation of excess nutrients in the overfed culture.

Optimization of culture conditions for high cell density proliferation of HL-60 human promyelocytic leukemia cells

1990 ◽  
Vol 97 (4) ◽  
pp. 639-647
Author(s):  
B. Schumpp ◽  
E.J. Schlaeger
Keyword(s):  
Tumor Necrosis Factor ◽  
Cell Density ◽  
Tumor Necrosis ◽  
Culture Conditions ◽  
High Density ◽  
Limiting Factors ◽  
Maximum Cell Density ◽  
Waste Products ◽  
Tumor Necrosis Factor Receptors ◽  
Necrosis Factor

The purpose of the present investigation was to optimize the culture conditions in suspension of the HL-60 cell line for high-density production. The optimized HL medium was a mixture of RPMI-1640, DMEM, HamF12 and IMDM media supplemented with transferrin, insulin, Primatone RL, Pluronic F68, ethanolamine and selenite. Under these conditions, whether serum was added or not, cells grew to up to 8 × 10(6) cells ml-1, which was at least three times higher than the maximum cell density usually described. Glucose and four amino acids: cystine, glutamine, methionine and serine, were highly consumed and disappeared quickly from the medium. Nutrient supply and metabolic end-product accumulation were the most probable growth-limiting factors. Different propagation systems were used to increase the cell density further. Cells were grown in dialysis tubing where relatively high levels of nutrients and low levels of waste products were maintained, leading to cell densities of 60 × 10(6) to 70 × 10(6) cells ml-1. A perfusion-culturing method with cell retention was found to be most effective for high-density production of HL-60 cells at a 21 as well as a 60 1 fermentor scale. Average concentrations of 40 × 10(6) to 50 × 10(6) cells ml-1 were achieved. Expression and distribution of both tumor necrosis factor receptors (55 and 75 × 10(3) Mr) on the surface of HL-60 cells were analysed as a control of the physiological integrity of the cells during the perfusion course; 1.3 micrograms of tumor necrosis factor receptors/10(10) cells was regularly expressed on the surface of HL-60 cells.

High density mammalian cell growth in Leibovitz bicarbonate-free medium: effects of fructose and galactose on culture biochemistry

1985 ◽  
Vol 78 (1) ◽  
pp. 173-189
Author(s):  
D. Barngrover ◽  
J. Thomas ◽  
W.G. Thilly
Keyword(s):  
Cell Growth ◽  
Mammalian Cell ◽  
Mdck Cells ◽  
High Density ◽  
Utilization Rate ◽  
Free Medium ◽  
Culture Ph ◽  
Base Form ◽  
Mammalian Cell Growth ◽  
Pyruvate Ratio

The most commonly used buffering system for mammalian cell cultures is a bicarbonate/CO2 system, which requires CO2 regulators and incubators to supply a constant level of CO2. As a replacement, Leibovitz developed a bicarbonate-free medium, L15, with relatively high levels of certain amino acids in the free base form. We found that a modified form of L15, containing 10 mM-fructose instead of galactose, supported high density growth of Vero and MDCK cells, with maintenance of a stable pH and lactate/pyruvate ratio. We report here investigations of Vero and MDCK cell growth and culture biochemistry at different concentrations of the two carbohydrates. The initial fructose concentration in the medium affected the eventual pH of the medium, the rate of production of lactic acid and ammonia, and the fructose utilization rate. The initial galactose concentration affected the growth rate but did not affect eventual culture pH, the rates of lactate and ammonia production, or the rate of its own utilization. Thus, Leibovitz' formula, modified to contain 10 mM-fructose, appears to yield satisfactory stability of culture pH and the lactate/pyruvate ratio. At all concentrations of galactose tested, the lactate/pyruvate ratio drifted out of the physiological range.

scholarly journals PDIA6 contributes to aerobic glycolysis and cancer progression in oral squamous cell carcinoma

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Ling Mao ◽  
Xiaoweng Wu ◽  
Zhengpeng Gong ◽  
Ming Yu ◽  
Zhi Huang
Keyword(s):  
Squamous Cell Carcinoma ◽  
Cell Growth ◽  
Oral Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Expression Pattern ◽  
Cancer Progression ◽  
Squamous Cell ◽  
Aerobic Glycolysis ◽  
Lactate Production ◽  
Control Group

Abstract Background/objective Accumulated evidence has demonstrated that aerobic glycolysis serves as a regulator of tumor cell growth, invasion, and angiogenesis. Herein, we explored the role of protein disulfide isomerase family 6 (PDIA6) in the aerobic glycolysis and the progression of oral squamous cell carcinoma (OSCC). Methods The expression pattern of PDIA6 in OSCC tissues was determined by qPCR and western blotting. Lentivirus and small interfering RNAs (siRNAs) were introduced into cells to upregulate and downregulate PDIA6 expression. CCK-8, flow cytometry, transwell, and xenotransplantation models were applied to detect cell proliferation, apoptosis, migration, invasion, and tumorigenesis, respectively. Results A high expression pattern of PDIA6 was observed in OSCC tissues, which was closely associated with lower overall survival and malignant clinical features in OSCC. Compared with the control group, overexpression of PDIA6 induced significant enhancements in cell growth, migration, invasiveness, and tumorigenesis and decreased cell apoptosis, while knockdown of PDIA6 caused opposite results. In addition, overexpression of PDIA6 increased glucose consumption, lactate production, and ATP level in OSCC cells. Conclusion This study demonstrated that PDIA6 expression was elevated in OSCC tissues, and overexpression of it promoted aerobic glycolysis and OSCC progression.

Amino acid uptake regulation by cell growth in cultured hepatocytes isolated from fetal and adult rats

1992 ◽  
Vol 12 (2) ◽  
pp. 135-141 ◽  
Author(s):  
S. Leoni ◽  
S. Spagnuolo ◽  
M. Massimi ◽  
F. Terenzi ◽  
L. Conti Devirgiliis
Keyword(s):  
Amino Acid ◽  
Cell Growth ◽  
Growth Inhibition ◽  
Cell Density ◽  
Amino Acid Uptake ◽  
Growth Stimulation ◽  
Acid Uptake ◽  
Adult Rats ◽  
Dependent Growth ◽  
Differentiation Stage

Amino acid uptake mediated by system A was studied in cultured fetal and adult hepatocytes, subjected to growth stimulation by EGF and insulin, or to growth inhibition by high cell density. The mitogenic stimulation induced a strong transport increase only in fetal cells, while the cell density-dependent growth inhibition, probably mediated by molecules present on adult hepatocyte membranes, provoked the decrease of amino acid uptake only in the adult cells. The results indicate that the different modulation of amino acid transport by cell growth is dependent on the age and the differentiation stage of hepatocytes.

A Comparison of CO2 and N2 Foaming Behaviors of PP in a Visualization System

2020 ◽  
Vol 35 (5) ◽  
pp. 503-517
Author(s):  
Q.-P. Guo ◽  
J. Wang ◽  
C. B. Park
Keyword(s):  
Cell Density ◽  
Nucleating Agent ◽  
Gas Content ◽  
Experimental Conditions ◽  
Maximum Cell Density ◽  
Visualization System ◽  
Cell Nucleation ◽  
Processing Strategies ◽  
Automotive Parts ◽  
Maximum Cell

Abstract Understanding of polypropylene (PP) foaming is critically important to reduce the weight of automotive parts. In this study, we used a batch foaming simulation system with visualization cell, to observe the foaming behaviors of PP that is blown with CO2 and N2 under various experimental conditions. We found that the nucleating agent content, initial temperature, pressure (i. e., gas content), and pressure drop rate during foaming have a significant effect on cell nucleation and cell growth. The cell density and the void fraction of PP foamed with CO2 and N2, respectively, were separately observed and compared. It was found that under the same experimental conditions, the maximum cell density of PP foamed with CO2 was higher than that of PP foamed with N2. However, the maximum cell density of PP foamed with CO2 was determined to be lower than that of PP foamed with N2, when the same gas mole numbers were employed. Based on the experimental results, optimum foaming conditions and effective processing strategies for PP-CO2 system are suggested.

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