Abstract

The human placenta and cord blood are rich in hematopoietic stem cells (HSCs), which give rise to all the blood cell types including the myeloid and lymphoid lineages. In addition, the placenta is also a source of mesenchymal stem cells (MSCs) that have a broad differentiation potential.¹ MSCs from fetal membranes and placental tissue are able to differentiate in vitro into multiple lineage cell types including osteoblasts, chondrocytes, myocytes, adipocytes, and endothelial cells. Hence, the potential applications of placenta and cord blood stem cells can be numerous.

Very little is known about stem cells in dolphins and marine mammals in general. We know that MSCs can be isolated from the adipose tissue of dolphins^2,3 that we assume have embryonic, fetal and adult stem cells like humans and the other studied mammals. But, in fact, dolphins have really unique adaptations and characteristics associated to the marine environment, so it would not come as a surprise that they could have special stem cell lineages or distinctive mechanisms of cell programming. For instance, we know that dolphin skin exhibits unique regenerating properties when wounded.^4-7 Reports of the survival after severe traumatic injury suggest that efficient healing of soft-tissue injury might be widespread among marine mammals.^8,9

We present preliminary data on the collection, isolation and characterization of stem cells from cord blood and placenta of the dolphin, Tursiops truncatus. The protocol has been created and refined through the sampling of 3 different females from 3 parks. Deliveries were successful and the placenta was collected from the water right after ejection, turned inside out to expose the fetal side, and sampled. Six ml of cord blood was collected from the umbilical veins, and HSCs were isolated and put to culture. HSCs at this stage were also frozen in cryopreserving media (CM) to test for the cryopreservation reliability. Cubes of 2 x 2 cm of placental tissue were excised from the fetal side in 2 to 4 regions with few/no veins and after enzymatic digestion and mechanical dissociation, the placental MSCs were put to culture. Small cubes were frozen in CM as well to test the potentiality of tissue cryopreservation for stem cells isolation. The cells will be characterized for expression of cell surface markers, embryonic stem cell gene expression and their differentiation ability. The phenotypic properties of the cells will be quantitatively described, and they will also be karyotyped.

If successful, the protocol developed could be used to create tissue banks for obtaining HSCs and MSCs in dolphins from aquatic parks for any future need. For therapeutic purposes, this could represent a great alternative to more invasive, time-consuming isolation/proliferation of autologous MSCs from a live dolphin. Moreover, the availability of a renewable source of dolphin stem cells will aid in the investigation of the unique and specific properties of HSCs and MSCs and all derived lines in dolphin vs. humans, leading to potential biomedical applications in regenerative medicine and treatment of various immune disorders.

Acknowledgements

The authors wish to thank the veterinarians Francesco Benaglia, Elena Campesi, Nicola Pussini and Matteo Sommer. We also thank the trainers and research staff at Oltremare, Acquario di Genova and Oceanografic parks who are daily committed to animal care and greatly contributed to the tissue sample retrieval.

* Presenting author

Literature Cited

1.  Matikainen T, Laine J. Placenta - an alternative source of stem cells. Toxicol Appl Pharmacol. 2005;207:544–549.

2.  Johnson SP, Catania JM, Harman RJ, Jensen ED. Adipose-derived stem cell collection and characterization in bottlenose dolphins (Tursiops truncatus). Stem Cells Dev. 2012;21(16):2949–2957.

3.  Griffeth RJ, García-Párraga D, Mellado-López M, Crespo-Picazo JL, Soriano-Navarro M, Martinez-Romero A, Moreno-Manzano V. Platelet-rich plasma and adipose-derived mesenchymal stem cells for regenerative medicine-associated treatments in bottlenose dolphins (Tursiops truncatus). PLoS One. 2014;9(9):e108439.

4.  Zasloff M. Observations on the remarkable (and mysterious) wound-healing process of the bottlenose dolphin. J Inv Derm. 2011;131:2503–2505.

5.  Bruce-Allen LJ, Geraci JR. Wound healing in the bottlenose dolphin. Can J Fish Aquat Sci. 1985;42:216–228.

6.  Corkeron PJ, Morris RJ, Bryden MM. A note on the healing of large wounds in the bottlenose dolphins. Aquat Mamm. 1987;13:96–98.

7.  Hicks BD, St. Aubin DJ, Geraci JR, Brown WR. Epidermal growth in the bottlenose dolphin, Tursiops truncatus. J Invest Dermatol. 1985;85(1):60–63.

8.  Van den Hoff J, Morrice MG. Sleeper shark (Somniosus antarcticus) and other bite wounds observed on southern elephant seals (Mirounga leonina) at Macquarie Island. Mar Mamm Sci. 2008;24:239–247.

9.  Bertilsson-Friedman P. Distribution and frequencies of shark-inflicted injuries to the endangered Hawaiian monk seal (Monachus schauinslandi). J Zool. 2006;268:361–368.