Umbilical cord and placenta blood banks in the form of frozen, amplifiable samples, could considerably shorten the wait for a transplant.

One of the major challenges facing cell biology is to recreate human tissues in vitro for reimplantation. Tissue cultures depend on a special category of cells: stem cells capable of multiplying - for self-renewal of the stock - and of differentiation to turn into specialised cells.

Research scientists and doctors are showing particular interest in blood stem cells or hematopoietic stem cells (HSC), found in the red bone marrow. These form every type of blood cell (red corpuscles, immune cells or platelets). In vitro cultivation of HSC could therefore provide a means to produce immune cells for patients with a bone marrow deficiency or red corpuscles to treat anaemia - or even to create a complete blood substitute for transfusions.

Multiplying without differentiation

All transplants require a sufficient supply of blood stem cells to reimplant and, above all, regenerate the tissue. However, as with all organs and blood, there are not enough donors. In the specific case of HSC, produced by the red marrow in the long bones - such as the femur or sternum - major surgery is needed to obtain the sample and this, in turn, has deterred many a compatible donor.

What is needed, therefore, is to amplify the rare supply of HSC available. There are two difficulties: stem cells are resting cells, which means they do not divide spontaneously and when they start proliferation and differentiation, they rapidly die. For this reason, the objective of the European Hematopoietic Bioreactor project was to amplify the HSC in vitro and achieve self-renewal, without differentiation or ageing. Eight research teams worked on this project for four years under the EU's Biotech 2 programme.

Umbilical cord blood

The first thing which the partners in this project did was to choose a source of hematopoietic stem cells which solves the donor problem: blood from the umbilical cord which is generally destroyed after birth. HSC from the umbilical cord are of higher quality, less prone to infection and, above all, less likely to be rejected by the recipient than bone marrow cells. 'We realised that the blood from a single cord, that is between 80 and 120 ml, contained as many blood stem cells as a normal sample of between 800 and 1200 ml from a donor,' explained project coordinator Dr Jacques Hatzfeld. 'Even these few drops of blood from a single cord can be amplified in vitro to produce enough stem cells for a graft on an adult.'

These precious cells are therefore recovered and cultured in bioreactors- a medium kept under constant control. Normally, however, cell division inhibitors keep the blood stem cells resting. The research teams countered this with molecules capable of blocking the inhibitors, thus clearing the way for the cells to enter the division phase. Next, they succeeded in slowing down the irreversible differentiation process. In this way, the HSC culture self-renews for over 100 days. Now the scientists are looking for ways of speeding up this self-renewal.

Placenta blood banks

Throughout Europe, particularly in Germany, France, Italy and Spain, umbilical cord and placenta blood banks are gradually being built up in the form of frozen samples, which, when the time comes, can then be amplified using the technique developed in this project. Before each sample is taken, the mother's blood is tested, to make sure that it is free of infections, and HLA-typed (1). The advantage of these HSC banks is that they considerably shorten the wait for a transplant. Once a compatible cell sample is found, it is available for the recipient immediately, compared with a wait of several weeks for a bone marrow transplant. In the longer term, doctors hope to build up a stock of compatible blood stem cells for each patient, from which they would produce the type of cell needed by the patient (lymphocytes, red corpuscles, etc.), depending on the case. It is already possible to control the results of HSC culture differentiation, by adding appropriate nutrients and growth factors.

Towards other tissue cultures

Another branch of this project is developing culture media containing no animal protein to reduce the risk of transmission of pathogens such as prions or viruses to humans. 'We have developed a medium containing no serumalbumin or transferin from cattle or human beings to carry the fatty acids and iron indispensable for any cell culture. We have replaced them by uncontaminated synthetic molecules performing the same function,' stated Dr Hatzfeld.

The blood stem cells used by these European researchers are, therefore, in a way a model which can be used in the future for other tissues in which industrial partners are already showing interest, such as liver, pancreas, bone or muscle. The way is therefore open to culturing stem cells for human tissues in Europe.

(1) HLA molecules: the 'identity card' carried by cells and deciding compatibility between donor and recipient.