Xylose-Based Maintenance Medium for Primate Pluripotent Stem Cells
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Solution Overview
Problem
Primate pluripotent stem cells, such as human ES cells and iPS cells, are challenging to maintain in undifferentiated and pluripotent states due to sensitivity to culture environment changes and low viability during cryopreservation, requiring complex and costly methods for long-term preservation and proliferation control.
Innovation Solution
A maintenance medium for primate pluripotent stem cells using xylose as a saccharide, substantially free of glucose, which maintains undifferentiation status and pluripotency, inhibits cell proliferation, and allows for short-term preservation without cryopreservation, utilizing xylose concentrations between 0.7 to 12.0 g/L in a DMEM/F12 medium.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If cryopreservation is performed for long-term preservation of primate pluripotent stem cells, then storage duration is extended, but cell viability decreases due to sensitivity to freezing and thawing
Solution Approach 1:
The invention changes the chemical composition parameters of the culture medium by replacing glucose with xylose and adjusting concentrations of bFGF and TGF-β3. This parameter change allows cells to be maintained in a dormant state with reduced metabolic activity, enabling long-term preservation without cryopreservation and maintaining high cell viability.
2Reliability
If feeder cells are used to maintain undifferentiation status and pluripotency, then cell properties are maintained, but culture process complexity increases
Solution Approach 1:
The invention extracts and isolates the essential factors (bFGF and TGF-β3) that maintain undifferentiation status from the complex feeder cell system. By using these purified factors in a defined medium with xylose, the invention eliminates the need for feeder cells while maintaining cell properties, thereby simplifying the culture process.
Solution Approach 2:
The invention introduces xylose as an intermediary substance that modifies cell metabolism and interacts with the growth factors bFGF and TGF-β3 to maintain undifferentiation status. This intermediary approach replaces the complex cell-cell interactions in feeder-based systems with a simplified chemical mediation system.
3Reliability
If substances for maintaining undifferentiation status (ascorbic acid, bFGF, TGF-β3) are added to the medium, then cell properties are maintained, but medium cost increases
Solution Approach 1:
The invention optimizes the concentration parameters of growth factors by combining xylose with reduced concentrations of bFGF and TGF-β3. This parameter optimization maintains effective pluripotency while reducing the total quantity of expensive substances required in the medium.
4Use of energy by moving object
If glucose is used as the saccharide in the culture medium, then cell metabolism is supported, but cell proliferation is excessive and properties change after certain passage numbers
Solution Approach 1:
The invention changes the saccharide parameter from glucose to xylose, which alters cell metabolism to a slower, more controlled rate. This metabolic modification allows cells to be maintained at stable properties for extended periods without excessive proliferation, reducing the need for frequent passaging and preserving cell identity.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The xylose-based medium effectively maintains the undifferentiation status and pluripotency of human iPS cells for extended periods, reduces the need for frequent medium changes, and lowers costs by utilizing abundant woody biomass resources, while allowing for controlled cell proliferation and preservation.
Implementation Method 1
Xylose is one of the constituent sugars of sugar chains, and plays an important role in vivo, such as in intercellular communication. However, in the field of culture cells, xylose has not been used as a saccharide in a medium because cells are considered not to be able to use xylose as energy.
Implementation Method 2
The present inventors found this time that in human iPS cells, when a medium in which glucose contained in a commercially available common medium for human iPS cells is replaced by xylose is used, the viability can be maintained and cell proliferation can be inhibited while the undifferentiation status and the pluripotency are maintained.
Data Source
AI summary
Provided are a maintenance medium for primate pluripotent stem cells, and a method for preserving and a method for controlling proliferation of primate pluripotent stem cells using the medium. The maintenance medium for human pluripotent stem cells according to the present invention includes xylose as a saccharide.