Figure 7

Recruitment and retention of pluripotent stem cells to infarcted myocardium in vitro. The autologous human induced pluripotent stem cells (ahiPSCs) were anchored to infarcted myocardia with the aid of heterospecific, tetravalent antibodies (htAbs). The data were validated with EDXS. (A) Efficacy of the htAbs to anchor ahiPSCs to cardiac muscle sarcomeres (ahiPSCs sarc) or endothelium (ahiPSCs endo). The htAbs were constructed from anti-SSEA-4, anti-TRA-1-60, anti- myosin, anti-α-actinin. For comparison, the efficacy of the htAbs to anchor the human cultured embryonic stem cells H1, H9 to the sarcomeres (hESC sarc) and to endothelium (hESC endo) and the PBMCs to the sarcomeres (hPBMC sarc) and endothelium (hPBMC endo) was determined. The measurements for each patient were conducted in triplicates. The data presented here are representative to all the samples studied. The statistical difference was accepted at P < .0003. (B) Retention of the anchored ahiPSCs onto the sarcomeres upon the day 1 of administration (ahiPSC sarc 1 d) was compared with that after 12 days (ahiPSC sarc 12 d). For reference, retention of the hESCs and the hPBMCs were quantified at the same time intervals. (C) Specificity of the htAbs to anchor the ahiPSCs to sarcomeres (ahiPSCs sarc) was tested by blocking the antigens on the ahiPSCs with non-specific tetravalent antibodies (anti-EGFRvIII, anti-EGFRvIV, anti-CEA, anti-PSMA) (ahiPSC sarc nsAb block) and with monospecific antibodies (anti-SSEA-4, anti-SSEA-3, anti-TRA-1-60, anti-TRA-1-81) (ahiPSCs sarc mAb block), or by omitting antibodies altogether (ahiPSCs no Ab). Blocking these biomarkers almost entirely eliminated anchoring of the ahiPSCs to the sarcomeres. The statistical significance was accepted at P < .0003. (D) Specificity of the htAbs to anchor the ahiPSCs onto the sarcomeres (ahiPSCs sarc) was measured by blocking antigens on the human cardiac muscle sarcomeres with non-specific tetravalent antibodies (anti-EGFRvIII, anti-EGFRvIV, anti-CEA, anti-PSMA) (ahiPSCs sarc nsAb block), with the monospecific antibodies (anti-myosin, anti-actin, anti-α-actinin, anti-titin) (ahiPSCs mAb block), or by omitting antibodies altogether (ahiPSCs no Ab). Blocking of these sarcomeric molecules almost entirely eliminated anchoring of the ahiPSCs to the sarcomeres. The statistical significance was accepted at P < 0.0003.