Pleiotrophin Is Highly Produced by Myeloma and Breast Cancer and Transdifferentiates Human Monocytes into Endothelial Cells That Are Incorporated into Tumoral Blood Vessels.
Authors: Chen Haiming , Campbell Richard A. , Li Mingjie , Gordon Melinda S. , Shalitin Dror , Wang Cathy S. , Berenson Ariana M. , Li Jennifer , Wong Hetty , Bonavida Benjamin , Berenson James R. 
Published: April 2020

Abstract
We have previously shown that multiple myeloma (MM) patients express pleiotrophin (PTN) and it is found at high levels in MM serum as well as PTN is a key factor in the transdifferentiation of monocytes into endothelial cells. We determined the level of PTN expression in myeloma and breast cancer and determined whether PTN produced by these tumor cells could induce endothelial cell expression in human monocytes. Both myeloma and breast cancer cells produced high levels of PTN and secreted this growth factor into the culture medium whereas normal bone marrow showed no expression of this protein. Next, MM cell lines, human bone marrow (BM) from MM patients or control subjects or breast cancer cells were cultured with CD14+ PBMCs using transwell culture plates coated with collagen I. CD14+ monocytes exposed to cells from MM cell lines or fresh BM or breast cancer cells showed expression of endothelial genes (Flk-1, Tie-2, CD144, and vWF) and lost expression of monocyte genes (c-fms). Induction of endothelial gene expression was blocked with an anti-PTN antibody. In contrast, CD14+ cells exposed to normal bone marrow as well as cell lines lacking PTN expression did not show endothelial gene expression. We determined whether human monocytes could be incorporated in vivo as vascular endothelium within human tumors that express PTN. Human myeloma LAGλ-1 cells which highly express and secrete PTN were mixed with THP1 monocytes transduced with the green fluorescent protein (GFP) gene and injected subcutaneously into SCID mice. Mice were sacrificed 6 weeks later and tumor was fixed and frozen sections. MM cells or THP1 monocytes alone did not demonstrate the presence of GFP+ blood vessels. Notably, GFP+ THP1 cells were found in blood vessels within the PTN-expressing LAGλ-1 tumor in animals injected with both cells together. When GFP+h2Kd- blood vessels were stained for anti-human and anti-mouse CD31, 60% of the endothelial cells stained positive for human CD31 and the remaining cells stained positive for mouse CD31 whereas none of these cells stained positive for both mouse and human markers. These results show that the blood vessels containing GFP+ cells do not result from fused cells. In addition, an anti-PTN antibody but not control IgG antibody blocks the incorporation of GFP+ cells into the vasculature of the LAGλ-1 tumors. Staining of serial sections with anti-Tie-2 and CD31 antibodies showed a similar distribution pattern. We further examined endothelial gene expression in these in vivo-generated samples using RT-PCR. The results showed that the THP1 monocytes or LAGλ-1 tumor cells alone did not express endothelial genes whereas THP1 monocytes mixed with PTN-expressing LAGλ-1 showed endothelial gene expression. This endothelial gene expression was blocked by anti-PTN antibody. These data show that hematologic and solid tumors through expression of PTN support new blood vessel formation by the transdifferentiation of monocytes into endothelial cells and provide a new potential target for inhibiting blood vessel formation in solid and liquid tumors.