Morris Hosseini

Human Umbilical Cord Perivascular (HUCPV) Cells: A Source of Mesenchymal Progenitors

We describe the isolation of a nonhematopoietic (CD45−, CD34−, SH2+, SH3+, Thy‐1+, CD44+) human umbilical cord perivascular (HUCPV) cell population. Each HUCPV cell harvest (2–5 × 106, depending on the length of cord available) gave rise to a morphologically homogeneous fibroblastic cell population, which expressed α‐actin, desmin, vimentin, and 3G5 (a pericyte marker) in culture. We determined the colony‐forming unit‐fibro‐blast (CFU‐F) frequency of primary HUCPV cells to be 1:333 and the doubling time, which was 60 hours at passage 0 (P0), decreased to 20 hours at P2. This resulted in a significant cell expansion, producing over 1010 HUCPV cells within 30 days of culture. Furthermore, HUCPV cells cultured in nonosteogenic conditions contained a subpopulation that exhibited a functional osteogenic phenotype and elaborated bone nodules. The frequency of this CFU‐osteogenic subpopulation at P1 was 2.6/105 CFU‐F, which increased to 7.5/105 CFU‐F at P2. Addition of osteogenic supplements to the culture medium resulted in these frequencies increasing to 1.2/104 and 1.3/104 CFU‐F, respectively, for P1 and P2. CFU‐O were not seen at P0 in either osteogenic or non‐osteogenic culture conditions, but P0 HUCPV cells did contain a 20% subpopulation that presented neither class I nor class II cell‐surface major histocompatibility complexes (MHC−/−). This population increased to 95% following passage and cryopreservation (P5). We conclude that, due to their rapid doubling time, high frequencies of CFU‐F and CFU‐O, and high MHC−/− phenotype, HUCPV cells represent a significant source of cells for allogeneic mesenchymal cell‐based therapies.

Effect of platelet releasate on bone cell migration and recruitment in vitro

The use of platelets and platelet products has become increasingly popular clinically as a means of accelerating endosseous wound healing. It is likely that growth factors released by activated platelets at the site of injury play a role in bone regeneration by stimulating the migration and proliferation of bone cells. In this study, a novel in vitro assay was developed to study the effects of platelet releasate (PR) collected from activated platelet concentrate on rat bone marrow–derived cells. Cultures of primary rat bone marrow cells were overlaid with a fibrin matrix, and the number of cells migrating within the three-dimensional matrix and the leading front of migration were quantified. The addition of PR to the top of the fibrin gels at different time points caused a 25% increase in the leading front of migration and a 3.5-fold increase in the number of migrating cells. Platelet releasate was also shown to have a mitogenic effect on bone cells in proliferation studies. Comparison between migration and proliferation data indicated that PR stimulates the initial recruitment of bone marrow cells to migration. This assay further allowed the determination that rat bone marrow cells are capable of exerting contractile forces on fibrin matrices and that matrix contraction is directly related to the migratory activity of cells. The results provide a potential mechanism to explain why biologically active platelet-derived factors enhance endosseous wound healing.