Work Package 6: Regenerative medicine

Professor Anna David, Professor Jan Deprest, Professor Paolo De Coppi

About this work package

This work package investigates the feasibility and efficacy of using stem cells and other regenerative therapies for the management of two types of fetal abnormalities: congenital haematopoietic disorders and myelomeningocele (MMC). Advances in our understanding of stem cells, prenatal diagnosis, prenatal imaging and fetal interventions have opened up new opportunities for the treatment of congenital abnormalities prior to the onset of symptoms.

Two main methodologies are evaluated:

  • Expansion and delivery of human fetal stem cells into fetal circulation
  • Application of tissue engineering approaches for the treatment of MMC in utero.

So far, the team has successfully achieved the expansion of various fetal stem cells types with efficient transplantation and engraftment into adult mice. Current work is on establishing a robust and safe protocol for the in utero delivery of cells into fetal circulation.

With respect to MMC, the team is investigating two aspects: patch repair and stem cell-based therapy. The team has optimised a protocol for fetoscopic spina bifida repair using a patch to seal the defect. They are also currently investigating the use of a novel tissue engineered patch and whether it would lead to better outcomes.

Work package tasks

Evaluate the feasibility, safety and efficacy of delivering cells, gene-constructs or other pharmacologically active treatments into the fetal circulation

Congenital haematopoietic disorders represent the earliest characterised diseases and are the most common genetic disorders. Recent studies in regenerative medicine have intensified the search for new stem cell sources with therapeutic potential. Fetal stem cells from cord blood, and amniotic fluid/ membrane have emerged as alternative sources of stem cells. Fetal stem cells are easy to isolate, more plastic than adult stem cells; have higher expansion and engraftment potential and lower immunogenicity. As a result, fetal stem cells are ideal candidates for cell and gene therapy applications. However, expansion of the fetal stem cells ex-vivo is key to increasing the effective cell dose of the graft for successful engraftment.

So far, the team has been successful in:

  • Isolation and expansion of cord blood stem cells and their long-term engraftment in fetal and adult immunocompromised NSG (NOD SCID il2rg-/-) mice
  • Isolation and expansion of amniotic fluid (AF) stem cells
  • Generation of AF-IPS (induced pluripotent stem) cells and their differentiation into haematopoietic cells for therapy

Preliminary work on in utero delivery of the cells into fetal circulation have been encouraging. Current work focuses on the in vivo delivery of stem cells into circulation to assess for teratoma formation and long-term engraftment potential in animal models. Further studies on in utero delivery would include donor-specific tolerance and survival.

Test the feasibility, safety and efficacy of treating MMC in utero using a single port access comparing early with standard timing of repair and using a tissue engineering approach

There are three areas of development in surgical MMC closure taking place currently:

  1. open fetal surgery is being converted to less invasive fetoscopic surgery, which inherently leads to a change in the way the MMC defect is closed and the need for  gas-amniodistension of the uterine cavity;
  2. while attempting to convert to fetoscopy, the number of ports to perform the surgery is being reduced from three or four, to two or even one;
  3. novel MMC defect coverage techniques are being developed, which may independently improve outcome, or may render the operation “simpler” hence amenable by fetoscopy through novel instruments and/or reduced number of ports.

We use the established sheep model, with creation of a lengthy spinal defect and myelotomy at 65 days through a hysterotomy. Since the principal outcome measure of this WP is the efficacy of the repair, we have first developed a comprehensive and sensitive panel of outcome measures for MMC-closure in the fetal sheep model. This includes measurement of reversal of hindbrain herniation on MR, cerebrospinal fluid leakage, conservation of somatosensory or motor evoked potentials at day 1 of live, and brain and medullary histology. These outcome measures have been first assessed in negative (normal) and positive (spina bifida) controls as well as those undergoing standard repair (treatment). We are now comparing outcomes of standard treatment against multiport fetoscopic repair using either Duragen dural and/or Integra skin patches, and a novel patch combining novel biomimetic nanocomposite materials with polymeric gels and tissue-engineered products, which are sutured in place. The effects of amnio-insufflation are being investigated simultaneously to determine an optimised protocol for the procedure.

Test the feasibility, safety and efficacy of treating MMC in utero using patch-augmented repair and cell-based therapy

MMC is a severe form of spina bifida wherein the spinal canal and the protective membranes push out on the back, in the form of a sac causing nerve damage, hydrocephalus, paralysis and bowel/urinary incontinence. The main goal of experimental strategies is to protect the exposed spinal cord prior to its damage and to avoid the devastating clinical manifestations associated with the disease. Repair using patches/scaffolds and fetal stem cells could augment current therapy for better patient outcomes. Stem-cell based strategies for spina bifida could replace/regenerate damaged cells and/or create a microenvironment to promote healing and protection of the damaged tissue.

As of now, the team has been working on:

  • Validation and testing of an anti-adhesive Polycaprolactone (PCL) patch using in vitro cytotoxic, degradation and leakage assays
  • Application of the PCL patch in a rabbit model of MMC and to study the outcome measures including survival, host-immune response, patch integration and adhesion formation – 80% survival rate was observed in the patch repair group
  • Isolation, expansion and characterization of amniotic fluid Mesenchymal stem cells from spina bifida babies for therapy

Current work focuses on comparing the migratory, neuro-regenerative, neuroprotective and angiogenesis capacities of AF-MSCs from normal and spina bifida babies.

Sign up for our studies

The Amniotic fluid, placental and fetal stem cells at birth study aims to find out whether we can grow stem cells from amniotic fluid, placental tissue and fetal fluid, and if proteins found in the fluid can be used to indicate a long-term outcome for problems such as kidney disease.

Find out more

Figure 1. The description is below the figure.

Figure 1. Potential application of fetal stem cells for Pre-natal and Post-natal Therapy: Fetal stem cells collected during gestation or at birth (1) can be purified and cultured ex-vivo (2). These cells be further expanded or manipulated (defective gene repair) (3) for autologous or allogeneic therapy pre-natally (5) or post-natally (6).

Key publications

Validation of the Fetal Lamb Model of Spina Bifida. Joyeux L, Engels AC, Van Der Merwe J, Aertsen M, Patel PA, Deprez M, Khatoun A, Pranpanus S, da Cunha MGMCM, De Vleeschauwer S, Parra J, Apelt K, Laughlin MM, Van Calenbergh F, Radaelli E, Deprest J. (2019). Scientific Reports, 9(1):9327. doi: 10.1038/s41598-019-45819-3. 

Learning curves of open and endoscopic fetal spina bifida closure: a systematic review and meta-analysis. Joyeux L, De Bie F, Danzer E, Russo FM, Javaux A, Peralta CFA, De Salles AAF, Pastuszka A, Olejek A, Van Mieghem T, De Coppi P, Moldenhauer J, Whitehead WE, Belfort MA, Lapa DA, Acacio GL, Devlieger R, Hirose S, Farmer DL, Van Calenbergh F, Adzick NS, Johnson MP, Deprest J.  (2019). Ultrasound in Obstetrics and Gynecology.  doi: 10.1002/uog.20389. [Epub ahead of print] Review 

Physiological effects of partial amniotic carbon dioxide insufflation with cold, dry vs heated, humidified gas in a sheep model. Amberg BJ, Hodges RJ, Kashyap AJ, Skinner SM, Rodgers KA, McGillick EV, Deprest JA, Hooper SB, Crossley KJ, DeKoninck PLJ. (2019). Ultrasound in Obstetrics and Gynecology. 53(3):340-347. doi: 10.1002/uog.20180

In utero therapy for congenital disorders using amniotic fluid stem cells. Subramaniam, S., Antoniadou, E., De Coppi, P., David, A. L.  (2018) Perinatal stem Cells: Research and Therapy, Book Chapter

Stem cell therapy in fetuses with spina bifida aperta: a systematic review and meta-analysis. Kunpalin, Y., Subramaniam, S., Perin, S., Bosteels, J., Ourselin, S., Deprest, J., De Coppi, P., David, A. L..  (Communicated to Prenatal Diagnosis)