Review
Hydroxyethylstarch in cryopreservation – Mechanisms, benefits and problems

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Abstract

As the progress of regenerative medicine places ever greater attention on cryopreservation of (stem) cells, tried and tested cryopreservation solutions deserve a second look. This article discusses the use of hydroxyethyl starch (HES) as a cryoprotectant. Charting carefully the recorded uses of HES as a cryoprotectant, in parallel to its further clinical use, indicates that some HES subtypes are a useful supplement to dimethysulfoxide (DMSO) in cryopreservation. However, we suggest that the most common admixture ratio of HES and DMSO in cryoprotectant solutions has been established by historical happenstance and requires further investigation and optimization.

Introduction

Demands for improved cell, tissue and organ storage are increasing as more and more products of regenerative medicine reach the clinic. The safety of cryopreservation of cell material is one of many emerging considerations in regenerative medicine. In addition use of cell-based assays for drug screening and safety testing raises questions as to what cryopreservation methods are preferable.

After briefly charting considerations in cryopreservation and cryoprotection, we will focus on one cryoprotectant factor in particular: We will consider the capacity of hydroxylethyl starch (HES) to act as a cryoprotectant, its use and its reported efficacy from the available data (which, although reaching back 40 years is still rather sparse) and consider the safety implications of using HES- in particular in comparison with the most common cryoprotectant compound dimethyl sulfoxide (DMSO).

Section snippets

Cryoprotection basics

Upon cooling with cryoprotectants, extra- and intracellular viscosities abruptly increase whereas thermal energy is decreased and not sufficient to enable chemical reactions. In this case all biological reactions are slowed down to a minimum that makes long term storage of cells, tissues and organs possible. The damages induced by cryopreservation involve many different cell compartments but the exact mechanisms are surprisingly poorly understood. In general, a distinction can be made between

HES structure

An alternative to simple sugars as cryoprotectants are complex carbohydrates made e.g. of glucose. One such example which has been used in cryopreservation is hydroxylethyl starch (HES), a synthetic modified polymer based on purified starch of corn or potatoes that has been modified by hydroxyethylation at carbon position C2, C3 or C6 [39], [40], [41] (see Fig. 1).

The physical, chemical and clinical properties of HES are depending on: [40] (see Fig. 2).

  • (i)

    The manufactured average molecular weight

Safety

A major disadvantage of cryoprotectants is their toxicity. Low-molecular penetrating CPAs can remove water from intracellular molecules, leading to unwanted interactions between proteins and to protein denaturation [72]. Thus, they have to be very carefully removed after cryopreservation, normally by centrifugation which has its own drawbacks. High-molecular cryoprotectants are usually better tolerated and do not necessarily need to be removed [70].

In pure form, all cryoprotectants are toxic to

Summary

Despite the list of successful cryopreservations of cells and tissues using HES, the mechanism of its action is weakly understood. This includes studies on the effect of molecular weight, degree of its molar substitution and purity on its success in cryopreservation. Given the general non-toxic profile of HES and the amounts likely involved in a cryopreserved clinical product, there seems ample scope to employ a wide range of HES with different MW, MS and degree of hydroxyethylation in order to

Declaration of interest

FV, NY and AS have in the past been funded by Serumwerke Bernburg AG, a manufacturer of HES.

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