previouslyWe shared the application of trehalose in human oligodendrocyte precursor cells（OPC）Cryoprotection of cell types such as adipose derived stem cells and mouse testicular germ cellsRelated studies have found that adding a certain amount of trehalose or using trehalose in frozen storage solution+Glycerol composite cryoprotectant can enhance the recovery rate of cells after long-term cryopreservation.

In this issue,We share an article by researchers from Milan, ItalyThe literature published in Stem Cell Research in 2018 examines how trehalose is used for cryopreservation of human pluripotent stem cells?

1. introduction

Cell products, including stem cells, have high clinical potential, and the entire stem cell market includes stem cell storage and preparation in the upper and middle reaches, as well as downstream clinical applications. According to a fully transparent market research institution（According to a report by Transparency Market Research, by the end of 2025, the total size of the stem cell market in China is expected to reach 270.5 billion US dollars, with a projected increase over the past 8 years

The compound annual growth rate of the stem cell market is expected to reach 13.8%.

In order to successfully apply cellular products, long-term low-temperature storage of cells is a technical difficulty that must be overcome, as low-temperature storage is a viable method for long-term preservation of cell vitality and biochemical functions.

In order to avoid damage to cell structure caused by intracellular freezing, some cryoprotectants have been launched. *The commonly used cell cryoprotectant is dimethyl sulfoxideDMSO is used in combination with fetal bovine serum or serum substitutes.

However, fetal bovine serum contains heterologous animal derived proteins, which may cause zoonotic infections with unknown pathogens. In addition,DMSO has cytotoxicity. According to reports, the toxicity of DMSO to tissues and cells depends on exposure time, temperature, and concentration. The degree of toxicity varies depending on the cell type, and there have been reports of adverse reactions in patients who re inject thawed cells without removing DMSO. In addition, it is known that DMSO affects the epigenome of mouse embryonic bodies by regulating the transcription levels of three DNA methyltransferases (Dnmts) and altering the genome-wide methylation profile, leading to uncontrolled differentiation of mouse stem cells. Therefore, dimethyl sulfoxide+fetal bovine serum is not suitable for clinical use, and there is an urgent need to develop efficient and non-toxic cell cryoprotectants.

Trehalose is a non reducing disaccharide that exists in various organisms that can survive in a fully dehydrated state, such as bacteria, yeast, slow-moving animals, and nematodes. Mammals do not produce trehalose, but trehalose is indeed an effective cryoprotectant that can reduce cell damage caused by ice crystal formation. Its protective effect is related to both osmotic effects and specific interactions between cell membrane phospholipids and unstable proteins, which can prevent damage and degeneration caused by dryness and oxidative stress.

Trehalose is not cytotoxic and has been effectively used for mouse germ cells, adult hematopoietic stem cells, mesenchymal stem cells, adipose derived stem cells, and human embryonic stem cellsCryostorage of different cell types such as hESCs and human induced pluripotent stem cells (hiPSCs). In addition, trehalose is also used for frozen storage of umbilical cord blood, bone marrow, human transplanted liver cells, and human pancreatic islets.

The main obstacle to the widespread use of trehalose in cell preservation is its difficulty in entering the interior of cells. Several methods have been applied in the past, such as osmotic shock, liposome delivery, thermal perforation, electroporation, microinjection, and genetic engineering, to help trehalose enter the cell interior. However, the above methods require very tedious operations, are laborious and time-consuming, and may lead to cell damage.

This study investigated the use of trehalose alone or in combination with ethylene glycol in the absence of dimethyl sulfoxide and fetal bovine serumFour different trehalose complex cryoprotectants were prepared by combining ethylene glycol or glycerol. Three different types of pluripotent stem cell lines were stored at low temperature and revived. Several key parameters were studied, including cell morphology, survival ability after thawing, expression level of pluripotent markers, genomic stability, endoplasmic reticulum (ER) homeostasis, DNA damage response, etc., to measure the low-temperature protective ability of different trehalose complex cryoprotectants on pluripotent stem cells.

Trehalose (for injection) (sterile)

1. Product Name: Trehalose (for injection) (sterile)

2. Chinese name: D (+) - trehalose dihydrate

3. English name: D (+) - Trehalose Dihydrate (for injection)

4. Chemical name: α - D-glucopyranosyl - α - D-glucopyranoside dihydrate

5. Model: Hipo-S

6. Content determination: 99.3%

7. CAS number: 6138-23-4

8. EINECS login number: 202-739-6

9. Registration number: F20190000452

10. DMF Number: 034401

11. Quality standards: comply with the pharmacopoeia standards of various countries (Chp, USP, JP, EP)

12. Molecular formula: C12H22O11 · 2H2O

13. Molecular weight: 378.33

14. Appearance: White crystalline powder

15. Specific rotation:+198.8 °

16. Acidity: 6.0

17. Chloride: compliant with regulations

18. Sulfate: compliant with regulations

19. Soluble starch: compliant with regulations

20. Burning residue: 0.04%

21. Heavy metals: comply with regulations

22、 水分:9.5%

23. Related substances: Total impurities 0.06%, glucose not detected, maltotriose 0.06%, * * unknown single impurities not detected

24. Aseptic: Compliant with regulations

25. Microbial limit: Aseptic

26. Inside bacteria:<0.1iu>

27. Nitrogen: compliant with regulations

28. Content determination: 99.3%

29. Solubility: soluble in water, soluble in hot ethanol, slightly soluble in methanol, insoluble in * ***

30. Storage: Store in a sealed, dark and dry place, and transport at room temperature.

31. Item number: 008001

32. Structural formula: