Regulatory T cells enhance mesenchymal stem cell survival and proliferation following autologous cotransplantation in ischemic myocardium Yifu Zhou,

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Presentation transcript:

Regulatory T cells enhance mesenchymal stem cell survival and proliferation following autologous cotransplantation in ischemic myocardium Yifu Zhou, MD, Avneesh K. Singh, PhD, Robert F. Hoyt, DVM, Suna Wang, MD, Zuxi Yu, MD, PhD, Timothy Hunt, BS, Bogdan Kindzelski, BS, Philip C. Corcoran, MD, Muhammad M. Mohiuddin, MD, Keith A. Horvath, MD The Journal of Thoracic and Cardiovascular Surgery Volume 148, Issue 3, Pages 1131-1137 (September 2014) DOI: 10.1016/j.jtcvs.2014.06.029 Copyright © 2014 Terms and Conditions

Figure 1 Diagram showing the experimental timeline from ameroid placement, mesenchymal stem cells and regulatory T cells injection, to the end of the experiment. MSC, Mesenchymal stem cell; LCX, left circumflex artery; LAD, left anterior descending artery. The Journal of Thoracic and Cardiovascular Surgery 2014 148, 1131-1137DOI: (10.1016/j.jtcvs.2014.06.029) Copyright © 2014 Terms and Conditions

Figure 2 Freshly isolated pig natural regulatory T cells were purified with flow cytometry cell sorting. Left, Gated lymphocytes. Center, CD4+ and CD25+ cells before sorting. Right, Both CD4+ and CD25+ regulatory T cells after sorting and ready for cotransplantation with mesenchymal stem cells. SSC, Side-scattered light; FSC, forward-scattered light. The Journal of Thoracic and Cardiovascular Surgery 2014 148, 1131-1137DOI: (10.1016/j.jtcvs.2014.06.029) Copyright © 2014 Terms and Conditions

Figure 3 Hematoxylin and eosin staining of a representative animal at 6 weeks after mesenchymal stem cells plus regulatory T cells injection showing spherical cell clusters with thin layers of capsules in frozen and paraffin-embedded sections. A, Arrow indicates the capsule around the cell clump (×100) in a frozen section. B, Higher magnification of (A) (×400). C, Three cell clusters were found inside myocardium (×100) in a paraffin-embedded section. D, Higher magnification of (C) (×200). The Journal of Thoracic and Cardiovascular Surgery 2014 148, 1131-1137DOI: (10.1016/j.jtcvs.2014.06.029) Copyright © 2014 Terms and Conditions

Figure 4 Hematoxylin and eosin staining of a representative animal at 6 weeks after mesenchymal stem cells injection without regulatory T cells (paraffin-embedded sections). A, Cell clusters were found in the border area of epi- and myocardium (×100). B, Higher magnification of (A) (×200). C, A cell cluster was found inside myocardium (×100). D, Higher magnification of (C) (×200). Arrows indicate the cell clusters. The Journal of Thoracic and Cardiovascular Surgery 2014 148, 1131-1137DOI: (10.1016/j.jtcvs.2014.06.029) Copyright © 2014 Terms and Conditions

Figure 5 Immunofluorescent staining of CD25 (green), CD90 (red), and 6-diamidino-2-phenylindole (blue) in frozen section at 6 weeks after mesenchymal stem cells plus regulatory T cells injection. A, CD25+ (regulatory T cells marker) cells were found inside the cell cluster with CD90+ (mesenchymal stem cell marker) cells (×200). B, Higher magnification of (A) (×400). Arrows indicate CD90+ cells and arrowheads indicate CD25+ cells. C, Stained with no first antibodies (control). D, Higher magnification of (C). The Journal of Thoracic and Cardiovascular Surgery 2014 148, 1131-1137DOI: (10.1016/j.jtcvs.2014.06.029) Copyright © 2014 Terms and Conditions

Figure 6 Immunofluorescent staining of Factor VIII (green), smooth muscle actin (red), and 6-diamidino-2-phenylindole (blue) in frozen section at 6 weeks after mesenchymal stem cells plus regulatory T cells injection. A, Increased capillary density suggested by Factor VIII+ and smooth muscle actin staining around the spherical cell cluster compared with (B) (no cluster area). C, A typical spherical cell cluster from another animal. D, Negative control (no antibodies were used). Arrows indicate cell clusters. Magnifications are ×100. The Journal of Thoracic and Cardiovascular Surgery 2014 148, 1131-1137DOI: (10.1016/j.jtcvs.2014.06.029) Copyright © 2014 Terms and Conditions