Characterisation of placental mesenchymal stromal cells and their role in cord blood transplantation.

Abstract

Cord blood transplantation (CBT) is an alternative to unrelated bone marrow transplantation in pediatric patients, while in adult patients the limited cell dose in cord blood (CB) unit results in delayed engraftment. To circumvent cell dose limitation, various methods have been investigated. Ex-vivo expansion of hematopoietic stem cells (HSC) is feasible but does not enhance engraftment due to HSC exhaustion. Use of double cord blood transplantation (DCBT) shows improved engraftment when compared to single unit transplantation, with median neutrophil engraftment at day 23 for recipients of DCBT, compared to 26-27 days for recipients of single cord blood transplantation (SCBT). However, engraftment is ultimately derived from single CB unit with reducing rates of chimerism seen up to day 100. The HSC share an intimate relationship with the BM microenvironment. Myeloablative conditioning using chemotherapy and radiotherapy may damage the microenvironment, which may contribute to delayed engraftment especially when the cell dose is limited. On the other hand mesenchymal stromal cells (MSC) could be used to restore this microenvironment. MSC are non-cycling cells having fibroblastic morphology, which express mesenchymal markers (CD73, CD105, CD90, CD29, and CD44), lack hematopoietic markers (CD45 and CD34) and differentiate into mesodermal lineages. MSC have been isolated from different tissues and show comparable characteristics to BM MSC. Recently, the placenta has been identified as a potential source of MSC and may have advantages to BM MSC due to a higher expansion potential and stronger immunosuppressive properties. This study has characterized cells obtained from the foetal aspect of the placenta and evaluated whether co-transplantation of placental MSC would enhance engraftment. Plastic adherent cells isolated from the placenta demonstrated typical characteristics of MSC. In 6 individual experiments, 4 cohorts of 24 NOD/SCID mice were evaluated. Cohort 1 received 5 × 10⁴ CD34⁺ cells from unit (U) 1 (SCBT); cohort 2 received 5 × 10⁴ CD34⁺ cells from U1+ 4 × 10⁴ MSC (SCBT+MSC); cohort 3 received 2.5 × 10⁴ CD34⁺ cells from U1+ 2.5 × 10⁴ CD34⁺ cells from U2 (DCBT); cohort 4 received 2.5 × 10⁴ CD34⁺ cells from U1+ 2.5× 10⁴ CD34⁺ cells from U2+ 4 × 10⁴ MSC (DCBT+MSC). Haemopoietic engraftment evaluated after 6-8 weeks was similar in recipients of SCBT and DCBT. MSC co-transplantation demonstrated enhanced engraftment in DCBT (51.8 ± 6.8% vs. 14.9 ± 6.5%; p=0.04) with an increased trend in SCBT (48.7 ± 7.7% vs. 17.5 ± 6.1%; p=0.07). In DCBT, co-transplantation of placental MSC reduced single CB unit dominance. Self-renewal capacity of engrafted HSC was assessed by serial transplantation in secondary recipients. Secondary recipients were infused with engrafted human cells from primary mice transplanted with or without MSC. In secondary transplantation of 17 evaluable mice, 13 engrafted at levels of 1-6.5%. Despite enhanced engraftment in primary mice, long-term engraftment capacity was unaltered with MSC co-transplantation. Furthermore, to study the potential mechanisms behind enhanced engraftment, eGFP transduced placental MSC and PKH-26 red labelled CB CD34⁺ cells were traced in live mice. Imaging studies showed MSC migrated to the pelvic region and improved CB CD34⁺ homing. Co-transplantation of placental MSC enhanced CB engraftment and may act by improving homing of CD34⁺ cells.