Mesenchymal Stromal Cells with Immunosuppressive Potential Can Be Mobilized Into Peripheral Blood by Electro-Acupuncture
Authors: Liu Lizhen , Yu Qin , Fu Shan , Dong Hui , Hu Kaimin , Zhang Lifei , Liu Senquan , Yu Xiaohong , Huang He 
Published: October 2019

Abstract
Abstract 4805

Background:
Mesenchymal stromal cells (MSCs) constitute a population of multipotential cells giving rise to adipocytes, osteoblasts and chondrocytes. Combining with their engraftment promoting capacity and immunosuppressive property, MSCs may be therapeutically useful for haematopoietic stem cell transplantation. A small number of MSCs can be mobilized into circulation by appropriate stimuli, such as hypoxia. However, there is little evidence for clinically useful methods for MSC mobilization. In this study, we used animal model to determine whether MSCs can be mobilized into peripheral blood (PB) by electro-acupuncture (EA), a traditional Chinese medical method.

Design and Methods:
Adult male Sprague-Dawley rats (200–220g) were randomly divided into there groups: EA-7 days, EA-14 days and control groups. For EA treatment, the rats were immobilized. A pair of stainless needles of 0.35mm diameter was inserted into points ‘Jizhong’ (GV6) and ‘Mingmen’ (DU4) and was then connected with output terminals of an EA apparatus. Alternating strings of dense-sparse frequencies were selected and the intensity was adjusted to induce slight twitch of the skin, with the intensity lasting for 30 min. Electro-acupuncture was applied to rats once a day for 7days or 14days. The control rats were immobilized for the same period without EA. To quantify the number of MSCs and evaluate mobilization efficiency, PB and bone barrow (BM) samples of each group were collected and colony-forming unit fibroblast (CFU-F) assays were performed. Mobilized PB derived MSCs were identified by immunophenotype and trilineage differentiation. Mixed lymphocyte reactions (MLR) were done to evaluate the immunosuppressive potential of mobilized MSCs and the cytokine levels TGF-β1, HGF and IL-10 in the supernatants of MSCs culture were measured by ELISA.

Results:
We found that MSCs can be mobilized into PB by electro-acupuncture. CFU-F frequency in rat PB was significantly increased after electro-acupuncture for 7days (8.20 ±1.48 vs.1.40 ±0.55 CFU-Fs per 3×106 cells) (p<0.05, n=5). PB CFU-F frequency increased to 12.4±1.82 per 3×106 cells in rats treated with electro-acupuncture for 14 days. However, no significant differences were observed in BM CFU-Fs among varies groups (P>0.05). Mobilized PB derived adherent cells were positive for CD90, CD29 and CD44, but negative for CD34 and CD45. After adipogenic, osteogenic, and chondrogenic induction, adherent cells from mobilized PB were positive for specific stains. In addition, they expressed mRNAs of Lpl and Pparg2 (adipocytic markers), Bglap and Runx2 (osteoblastic markers), and Col2a1 and Col10a1 (chondrocytic markers). These results showed that mobilized PB-derived cells could differentiate into adipocyte, osteoblast, and chondrocyte, which indicated that they are bona fide MSCs. The levels of immunosuppressive cytokines TGF-β1, HGF and IL-10 in the supernatants of PB-MSC culture were similar as BM MSCs. Results of MLR showed that mobilized PB derived MSCs inhibited T lymphocyte proliferation.

Conclusion:
Taken together, these data revealed, for the first time to the best of our knowledge, that multipotential MSCs can be mobilized by electro-acupuncture. Our study provides a clinical useful method to mobilize MSCs with immunosuppressive potential and highlight a novel insight into the mechanisms of electro-acupuncture therapy.

Disclosures:
No relevant conflicts of interest to declare.