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Glucose utilization, pH reduction and density dependent inhibition in cultures of chick embryo fibroblasts
Insights
Chick embryo cells show reduced multiplication at pH 6.9, especially before confluency. Glucose metabolism to lactic acid and pH changes do not fully explain this density-dependent growth inhibition.
Area of Science:
- Cell Biology
- Biochemistry
Background:
- Cell multiplication is influenced by environmental factors like pH and nutrient availability.
- Density-dependent inhibition (DDI) limits cell proliferation in crowded cultures.
Purpose of the Study:
- To investigate the impact of pH and glucose metabolism on chick embryo cell multiplication.
- To determine the role of lactic acid production and glucose depletion in DDI.
Main Methods:
- Culturing chick embryo cells at different pH levels (7.4 and 6.9).
- Measuring glucose uptake, lactic acid production, and cell multiplication rates.
- Analyzing the relationship between glucose concentration and cell growth.
Main Results:
- Cell multiplication rate was only slightly lower at pH 6.9 compared to pH 7.4, but showed marked reduction before confluency.
- 80-90% of glucose was converted to lactic acid, significantly lowering medium pH.
- Neither lactic acid accumulation nor pH reduction fully explained DDI.
- Glucose depletion can contribute to growth inhibition in very crowded cultures.
Conclusions:
- Lactic acid production and associated pH decrease are not the primary drivers of DDI in chick embryo cells.
- While glycolysis is not limited by glucose uptake at physiological concentrations, glucose depletion can inhibit growth in high-density cultures.
Abstract:
The multiplication rate of sparse cultures of chick embryo cells is only slightly lower at pH 6.9 than at pH 7.4. There is, however, a marked reduction in the multiplication rate of the pH 6.9 cultures before they reach confluency. Cultures at pH 7.4 continue to multiply beyond confluency with only a slight decrease in the multiplication rate. Eighty to ninety percent of the glucose taken up by the cells growing at each pH is converted to lactic acid which is released into the medium. Metabolic reduction in pH of the medium is almost entirely accounted for by the amount of lactic acid produced by the cells. Neither the intracellular nor extracellular accumulation of lactic acid nor the accompanying reduction in pH is sufficient to explain density dependent inhibition of the rate of multiplication of chick cells. The rate of lactic acid production and the multiplication rate of chick cells are independent of glucose concentration in the range of 2--16 mM. In view of the kinetic parameters for the uptake of glucose, this shows that glycolysis is not limited by the rate of glucose uptake and that depletion of glucose from the medium cannot account for the onset of density dependent inhibition of multiplication. However, when cells reach very high population densities, conventional glucose concentrations of 5 mM can be depleted overnight by chick cells. Since the multiplication rate of cells is dependent on glucose concentration when it falls below 2 mM, depletion of glucose may cause some growth inhibition in crowded cultures supplied with conventional medium.