Abstract 4799

We have previously shown that microvesicles derived from irradiated murine lung cells enter murine bone marrow cells and induce expression of the pulmonary epithelial cell-specific mRNAs Surfactants A-D (Sp-A-D), Aquaporin-5 (Aq-5) and Clara Cell Specific Protein (CCSP). The early mRNA increases are due to direct delivery of mRNA and due to the transfer of a transcription factor to marrow cells (Aliotta JM, et al, Exp Hematol, 2010). The modulated marrow cells produce pulmonary epithelial cell-specific protein and have an enhanced ability to convert into pulmonary epithelial cells after transplantation into lethally-irradiated mice. In the present studies, we evaluated the stability of pulmonary epithelial cell-specific mRNA expression in cytokine-stimulated liquid culture and after in vivo transplantation into lethally-irradiated mice. Whole bone marrow cells isolated from C57BL/6 mice were cultured for seven days opposite C57BL/6 lung fragments, but separated from them by a 0.4 micron cell-impermeable membrane. Marrow cells were then either infused into lethally-irradiated C57BL/6 mice or established in culture with DMEM supplemented with IL-3, IL-6, IL-11 and steel factor. Engrafted mice were sacrificed six weeks after transplantation and real-time RT-PCR was carried out on marrow, liver, spleen and thymus tissue. Expression of Sp-A, B and D, CCSP and Aq-5 were variably seen in all tissues and increases over control ranged from 12-fold to 58-fold. Liquid culture was analyzed every two weeks out to 12 weeks and high levels of expression of different pulmonary epithelial cell-specific mRNAs and proteins (by western blot and Immunohistochemical staining) were observed. In order to determine the mechanism for the increased expression in culture, we established co-cultures of Fischer-344 rat lung opposite C57BL/6 mouse bone marrow cells and, using species-specific primers, determined whether the increased pulmonary epithelial cell-specific mRNA expression at each time point of secondary cytokine-supported culture was rat or mouse-specific. In every instance, the pulmonary epithelial cell-specific mRNA which was detected was mouse-specific, not rat-specific, and thus the long-term expression of lung-specific mRNA in mouse marrow was due to induced transcription by the mouse marrow cells. These data indicate that lung-derived microvesicles induce long-term stable epigenetic changes in marrow cells and suggests that the genetic changes induced by tissue microvesicles in neighboring cells may be biologically important.

Disclosures:

No relevant conflicts of interest to declare.

Author notes

*

Asterisk with author names denotes non-ASH members.

Sign in via your Institution