Maria De Ioanni

Mesenchymal cells recruit and regulate T regulatory cells

OBJECTIVE Despite much investigation into T regulatory cells (Tregs), little is known about the mechanism controlling their recruitment and function. Because multipotent mesenchymal stromal cells (MSCs) exert an immune regulatory function and suppress T-cell proliferation, this in vitro study investigated their role in Treg recruitment and function. MATERIALS AND METHODS Human MSCs and different T cell populations (CD3(+), CD3(+)/CD45RA(+), CD3(+)/CD45RO(+), CD4(+)/CD25(+), CD4(+)/CD25(+)/CD45RO(+), CD4(+)/CD25(+)/CD45RA(+)) from healthy donors were cocultured for up to 15 days. Harvested lymphocytes were analyzed by flow cytometry and FoxP3 and CD127 expressions were measured by real-time polymerase chain reaction. Their regulatory activity was assessed. RESULTS We demonstrate MSC recruit Tregs from a fraction of CD3(+) and from immunoselected CD3(+)/CD45RA(+) and CD3(+)/CD45RO(+) fractions. After culture with MSCs both immunoselected fractions registered increases in the CD4(+)/CD25(bright)/FoxP3 subset and CD127 expression was downregulated. When purified Treg populations (CD4/CD25(+), CD4/CD25(+)/CD45RA(+), and CD4/CD25(+)/CD45RO(+)) are used in MSC cocultures, they maintain FoxP3 expression and CD127 expression is downregulated. Treg suppressive capacity was maintained in Treg populations that were layered on MSC for up to 15 days while control Tregs lost all suppressive activity after 5 days culture. CONCLUSIONS In conclusion, our study demonstrates that MSCs recruit, regulate, and maintain T-regulatory phenotype and function over time.

Unique human CD133+ leukemia cell line and its modulation towards a mesenchymal phenotype by FGF2 and TGFβ1†

Immunological features of GM‐490 cells, a new blood cell line from a patient with acute lymphoblastic leukemia, included lack of CD34, CD38, CD45, CD14, HLA‐DR, and lymphoid and myeloid markers and expression of CD29, CD36, CD44, CD54, CD71, CD105, and CD133. Molecular analysis indicated CD45 gene expression was absent but CD34 mRNA was present. GM‐490 cells constitutively produced fibronectin (FN), type III and traces of type I collagen, collagenases, glycosaminoglycans (GAG) and biglycan and betaglycan proteoglycans (PG) as well as FGF2 and TGFβ1. When FGF2 and/or TGFβ1 were added to cells in vitro, they stimulated cell proliferation and differently modulated matrix production and growth factor receptor expression. Reverse transcription‐polymerase chain reaction (RT‐PCR) detection of transcripts encoding for osteocalcin and RUNX2 suggests GM‐490 cells differentiate towards the osteoblast pathway. GM‐490 cells expressed the low affinity nerve growth factor receptor (p75LNGFR), a somatic stem cell marker that is not detected in hematopoietic cells, leading to the hypothesis that GM‐490 has mesenchymal stem cell properties. The reciprocal modulating effects of FGF2 and TGFβ1 on each others receptors make the GM‐490 cell line a new model for investigating the relationship between these growth factors and their receptors in autocrine loops which are believed to sustain the malignant clone in hematological diseases.